IE51393B1

IE51393B1 - Sulfamic acid derivatives

Info

Publication number: IE51393B1
Application number: IE2297/86A
Authority: IE
Original assignee: Squibb & Sons Inc
Priority date: 1980-02-07
Filing date: 1981-02-06
Publication date: 1986-12-24
Also published as: MY8600179A; JPH0670006B2; IE862297L; NO170015B; PT72465B; DD156180A5; GR74151B; PT72465A; ES8205397A1; GB2071650A; GB2139618B; NO170015C; IT8119587A0; AU569407B2; NO860225L; JPH0623188B2; GB2139618A; YU31981A; NZ196202A; DE3104145C2

Abstract

This invention concerns intermediates for use in preparing beta -lactam antibiotics. More specifically the invention provides a compound of the formula wherein R1 is hydrogen or acyl or the group R1-NH- is protected amino; R3 and R4 are the same or different and each is hydrogen, alkyl, cycloalkyl, phenyl or substituted phenyl, or one of R3 and R4 is hydrogen and the other is alkoxycarbonyl, alken-1-yl, alkyn-1-yl, 2-phenylethenyl or 2-phenylethynyl; V is a leaving group; and M<+> is hydrogen or a cation. It also provides similar compounds where R1 is a protecting group.

Description

This invention concerns a novel family of β-lactam antibiotics, and more specifically certain sulfamic acid derivatives used as intermediates in their preparation.

It has been discovered that the β-lactam nucleus can be biologically activated by a sulfonic acid salt substituent attached to the nitrogen atom in the nucleus. β-Lactams having a sulfonic acid salt (including inner salt) substituent in the 1-position and an acylamino substituent in the 3-position exhibit activity against a range of gram-negative and gram-positive bacteria. Such β-lactams are claimed in our copending Irish Patent Application No. 230/81.

The present application is particularly concerned with an intermediate for use in preparing such β-lactams, the intermediate being a compound of the formula:20 V wherein R1 is hydrogen or acyl or the group R^-NH- is protected amino; Rg and R4 are the same or different and each is hydrogen, alkyl, cycloalkyl, phenyl or substituted phenyl, or one of Rg and R^ is hydrogen and the other is alkoxycarbonyl, 25 alken-1-yl, alkyn-1-yl, 2-phenylethenyl or 2-phenylethynyl; V is a leaving group; and M+ is hydrogen or a cation.

Preferably R^NH is protected amino and such a compound may be prepared by sulfonating the corresponding amide (see the description of compounds of formulae XX and XX1 below).

The preferred members of the novel family of β-lactam antibiotics are those encompassed by the formula R. -NH-C 1 I In addition to the above described B-lactams having a sulfonic acid salt substituent in the 1-position and an aeylamino substituent in the 3-position, this invention also concerns β-lactams having a sulfonic acid salt (including inner salt) substituent in the 1-position and an amino substituent in the 3-position.

The preferred compounds of this type have the formula la Rj R4 C-R These compounds are intermediates useful for the preparation of corresponding 3- aeylamino compounds.

As used in formulas I and la, and throughout the specification, the symbols are as defined below.

R^ is acyl; Rj is hydrogen dr alkoxy of 1 to 4 carbons; R, and R. are the same or different and each 3 4 is hydrogen, alkyl, cycloalkyl, phenyl or substituted phenyl, or one of Rj and R^ is hydrogen and the other is alkoxyearbonyl, alken-l-yl, alkyn-l-yl, 2-phenylethenyl or 2-phenylethynyl.

M* is hydrogen, or a cation. -3The terms alkyl and alkoxy refer to both straight and branched chain groups. Those groups having 1 to 10 carbon atoms are preferred.

The terms eycloalkyl and cycloalkenyl refer to eycloalkyl and cycloalkenyl groups having 3,4,5,6 or 7 carbon atoms.

The term 'hlkenyl refers to both straight and branched chain groups. Those groups having 2 to 10 carbon atoms are preferred.

The term halogen refers to fluorine, chlorine, bromine and iodine.

The tern substituted phenyl refers to a phenyl group substituted with 1, 2 or 3 amino, halogen, hydroxyl, trifluoromethyl and lower alkyl or alkoxy groups.

The term protected carboxyl refers to a carboxyl group which has been esterified with a conventional -ester protecting group.

These groups are well known in the art; see, for example, U.S. Patent Specification No. 4.144.331 issued March 13, 1979. The preferred protected carboxyl groups are benzyl, benzhydryl and t-butyl esters.

The term acyl includes all organic radicals derived from an organic acid (i.e., a carboxylic acid) by removal of the hydroxyl group. Certain acyl groups are, of course, preferred, but this preference should not be viewed as a limitation of the scope of this invention. Exemplary acyl groups are those acyl groups which have been used in the past to acylate β-lactam antibiotics including 6-aminopenicillanic acid and derivatives and 7-aminocephalosporanic acid and derivatives; see, for example, Cephalosporins and Penicillins, edited by Flynn, Academic Press (1972), German Offenlegungsschrift 2,716,677 published October 10, 1978, Belgian Patent Specification No. 867,994, published December 11, 1978, U.S. Patent Specification No. 4,152,432, issued May 1, 1979, U.S. Patent Specification No. 3,971,778, issued July 27, 1976, U.S. Patent Specification No. 4,172,199, issued October 23, 1979, and British Patent Specification No. 1,348,894, published March 27, 1974.

The portions of these references describing various acyl groups are incorporated herein by reference.

The following list of acyl groups is presented to further exemplify the term acyl"; it should not be regarded as limiting that term. Exemplary acyl groups are: wherein Rj is alkyl; cycloalkyl; alkoxy; alkenyl; cycloalkenyl; cyclohexadienyl; or alkyl or alkenyl substituted with one or more halogen, cyano, nitro, amino, mercapto, alkylthio, or cyanomethyl thio groups. (b) Carbocyclic aromatic groups having the formula II Ri -5•0-CH0 II -C-, wherein n is 0, 1, 2 or 3; R-, R?, and R_ each ο 3.5 is independently hydrogen, halogen, hydroxyl, nitro, amino, cyano, trifluoromethyl, alkyl of 1 to 4 carbon atoms, alkoxy of 1 to 4 carbon atoms or aminomethyl; and Rg is amino, hydroxyl, a carboxyl salt, protected carboxyl, formyloxy, 2q a sulfo salt, a sulfoamino salt, azido, halogen, hydrazino, alkylhydrazino, phenylhydrazino, or [(alkylthio)thioxomethyl]thio.

Preferred carbocyclic aromatic acyl groups include those having the formula o-< -ch2-c513 9 3 a carboxyl •alt or sulfo salt) (Rg is preferably and (Rg is preferably a carboxyl salt or sulfo salt). (c) Heteroaromatic groups having the formula 0 RgffCH-&- , R9 O RlrfOCHg-C- , II g or 0 K lj RKj-c-ά- , wherein n is 0, 1, 2 or 3; Rg is as defined above; and R^is a substituted or unsubstituted -, 6-or 7-membered heterocyclic rinq containing 1, 2, 3 or 4 (preferably 1 or 2) nitrogen, oxygen and sulfur atoms. Exemplary heterocyclic 513/3 -7rings are thienyl, furyl, pyrrolyl, pyridinyl, pyrazinyl, thiazolyl, morpholinyl, pyrimidinyl and tetrazolyl. Exemplary substituents are halogen, hydroxyl, nitro, amino, cyano, trifluoro5 methyl, alkyl of 1 to 4 carbon atoms, or alkoxy of 1 to 4 carbon atoms.

Preferred heteroaromatic acyl groups include those groups of the above formulas wherein R^gis 2-amino-4-thiazolyl, 2-amino-5-halo10 4-thiazolyl, 4-aminopyrimidin-2-yl, 5-amino-l,2,4thiadiazol-5-yl, 2-thienyl or 2-furanyl. (d) [ ί (4-Substituted-2,3-dioxo-l-piperazinyl)carbonyl]amino]arylacetyl groups having the formula r~\ O 0 ll ll -C-CH-NH-C-N wherein Rj^ is an aromatic group (including carbocyclic aromatics such as those of the formula R, and heteroaromatics as included within the definition of Rj^; and R^2 is alkyl, substituted alkyl (wherein the alkyl group is substituted with one or more halogen, cyano, nitro, amino or mercapto groups), arylmethyleneamino (i.e., -N«CH-R-£j_ wherein is as defined above), aryl carbonyl amino 11 (l . e., — NH—C—wherein R11 is as defined above) or alkylcarbonylamino. -8Preferred [ I (4-substituted-2,3-dioxo-lpiperazinyl, carbonylJ aminoJ arylacetyl groups include those wherein R12 is ethyl, phenylmethyleneamino or 2-furylmethyleneamino. (e) (Substituted oxyimino)arylacetyl groups having the formula ll -c-on-o-r13 wherein Rj^is as defined above and R13 is hydrogen, alkyl, cycloalkyl, alkylaminocarbonyl, arylaminoO carbonyl (i.e., -ίί-ΝΗ-Κχχ wherein R^is as defined above) or substituted alkyl (wherein the alkyl group is substituted with 1 or more halogen, cyano, nitro, amino, mercapto, alkylthio, aromatic group (as defined by R]J , carboxyl (including salts thereof), amido, alkoxycarbonyl, phenylmethoxycarbonyl, diphenylmethoxycarbonyl, hydroxyalkoxyphosphinyl, dihydroxyphosphinyl, hydroxy (phenylmethoxy)phosphinyl, or dialkoxyphosphinyl substituents).

Preferred (substituted oxyimino,arylacetyl groups include those wherein R]jis 2-amino-4-thiazolyl. Also preferred are those groups wherein R33 is methyl, ethyl, carboxymethyl, or 2-carboxyisopropyl. (f, (Acylamino)arylacetyl groups having the formula wherein Rjjis ·· defined above and R^4 is -9(CH,) (cyanoalkyl) amino, -0-, amino, alkylamino, amido, alkylamido, (cyanoalkyl) Preferred (aeylamino)arylacetyl groups of the above formula include those groups wherein R^ is amino, or amido. Also preferred are those groups wherein R^is phenyl or 2-thienyl (g) (([3-Substituted-2-oxo-l-imidazolidinyl]carbonyl]amino]arylacetyl groups having the formula 0 II 0 θ -C II ll / \ -C-CH-NH-C-N N-R.' I I 15 CHj-CHj -10wherein i·8 38 defined above and R15 is hydrogen, alkylsulfonyl, arylmethyleneamino (i.e., -N»CH-R[j wherein R^is as defined above), II -cr16 ("herein Rj_g is hydrogen, alkyl or halogen substituted alkyl), aromatic group (as defined by R^above), alkyl or substituted alkyl (wherein the alkyl group is substituted with one or more halogen,cyano, nitro, amino or mereapto groups).

Preferred II3-substituted-2-oxo-l-iraidazolidinyl]carbonyl]amino]arylacetyl groups of the above formula include those wherein Ruis phenyl or 2-thienyl. Also preferred are those groups wherein R15 is hydrogen, methylsulfonyl, phenylmethyl eneamino or 2-£urylmethyleneamino.

The term cation, as used throughout the specification, refers to any positively charged θ β atom or group of atoms. The -SO^M " substituent 20 on the nitrogen atom of the β-lactams of this invention encompasses all sulfonic acid salts. Pharmaceutically acceptable salts are, of course, preferred, although other salts are also useful in purifying the products of this invention or as intermediates for the preparation of pharmaceutically acceptable salts. The cationic portion of the sulfonic acid salts of this invention can be obtained from either organic or inorganic bases. Such cationic portion includes, but is not limited to, the following ions: ammonium; substituted ammonium, such as alkylammonium (e.g., tetra-n-butylammonium, referred to hereinafter as tetrabutylammonium); alkali metal, such as lithium, sodium and potassium; alkaline earth metal, such as calcium and magnesium; -115 13 3 3 pyridinium; dicyclohexylammonium; hydrabaminium; benzathinium; N-methyl-D-glucaminium.

As set forth in formula I, and in the definitions following fonnula I, M® can be hydrogen.

As is described hereinafter, the e-lactams can be prepared by synthetic means. The nonalkoxylated 4-unsubstitubed β-lactams of formula I, i.e., those compounds of formula I wherein Rj, R^ and R^ are hydrogen can be prepared using 6-aminopenicillanic acid or a 6-acylaminopenicillanic acid as a starting material. The Β-lactams of formula I wherein is alkoxy can be prepared from the corresponding non-alkoxylated B-lactam.

Some of the compounds of this invention may be crystallized or recrystallized from solvents containing water. In these cases water of hydration may be formed. This invention contemplates stoichiometric hydrates as well- as compounds containing variable amounts of water that may be produced by processes such as lyophilization.

Some of the β-lactams of fonnula I have also been prepared by biological means. Culti25 vation of a strain of the microorganism Chromobacterium violaceum SC 11,378, yields a salt of (R)-3-(acetylamino)-3-methoxy-2-oxo-lazetidinesulfonic acid. Cultivation of various acetic acid bacteria e.g., Gluconobacter species SC 11,435, yields a salt of (R)-3-.((N-(D-y-glutamyl)D-alanyl ] aminop3-methoxy-2-oxo-l-azetidinesulfonic acid -12β-Lactams having a sulfonic acid salt substituent in the 1-position and an amino or acylamino substituent in the 3-position contain at least one chiral center the carbon atom (in the 3-position of the β-lactam nucleus) to which the amino or acylamino substituent is attached. This invention is directed to those β-lactams which have been described above, wherein the stereochemistry at the chiral center in the 3-position of the β-lactam nucleus is the same as the configuration at the carbon atom in the 6-position of naturally occurring penicillins (e.g., penicillin G) and as the configuration at the carbon atom in the 7-position of naturally occurring cephamycins (e.g., cephamycin C). with respect to the preferred β-lactams of formulas I and la, the structural formulas have been drawn to show the stereochemistry at the chiral center in the 3-position. Because of the nomenclature convention, those compounds of formulas I and Ia wherein R? Is hydrogen have the S configuration and those compounds of formulas I and Ia wherein R? is alkoxy have the R configuration.

Also included are racemic mixtures which contain the above-described f5-lactams. -13β-Lactams having a sulfonic acid salt substituent in the 1-position of the β-lactam nucleus and an amino or aeylamino substituent in the 3-position of the β-lactam nucleus have activity against a range of gram-negative and gram-positive organisms. The sulfonic acid salt substituent is essential to the activity of the compounds of this invention. The compounds wherein Rg and/or R4 are hydrogen or alkyl, especially methyl, exhibit especially useful activity.

The novel β-lactams can be used as agents to combat bacterial infections (including urinary tract infections and respiratory infections) in mammalian species, such as domesticated animals (e.g., dogs, cats, cows, horses, and the like) and humans.

For combating bacterial infections in mammals the β-lactam can be administered to a mammal in need thereof in an amount of about 1.4 mg/kg/day to about 350 mg/kg/day, perferably about 14 mg/kg/day to about 100 mg/kg/day. All · modes of administration which have been used in the past to deliver penicillins and cephalosporins to the site of the infection are also contemplated for use with the novel family of β-lactams.

Such methods of administration include oral, intravenous, intramuscular and as a suppository -14The novel β-lactam products are generally prepared by the introduction of a sulfonic acid substituent (a sulfo group -SOj-J onto the nitrogen atom in the 1-position of the Β-lactam nucleus. This sulfonation reaction is readily effected by treating the β-lactam with a sulfur trioxide complex or with an equivalent sulfonating reagent such as a chlorosulfonate.

The sulfur trioxide complexes most commonly used are pyridine-sulfur trioxide; lutidine-sulfur trioxide; dimethylformamide-sulfur trioxide; and picoline-sulfur trioxide. Instead of using a pre-formed complex, the complex can be formed in situ, e.g., using chlorosulfonyl-trimethylsilyl ester and pyridine as reagents. Alternatively, sulfonation can be effected by way of an intermediate compound as for exanple first silylating the nitrogen atom of the β-lactam nucleus and then subjecting the silated compound to a silyl interchange reaction with trimethylsilylchlorosulfonate or a similar reagent. Exemplary silylating agents are monosilyltri£luoroacetamide, trimethylsilylchloride/triethylamine and bis-trimethylsilyl trifluoroacetamide.

Generally, the sulfonation reaction is carried out in the presence of an organic solvent such as pyridine of amixture of organic solvents, preferably a mixture of a polar solvent such as dimethylformamide and a halogenated hydrocarbon such as dichloromethane^.

The product initially formed in the sulfonation reaction is a salt of the sulfonated β-lactam.

When pyridine-sulfur trioxide is the sulfonating complex the product initially formed is the β-lactam-sulfonated pyridinium salt of the sulfonated β-lactam wherein M+ in the formula below is the pyridinium ion: J-n-so,m+ 3 -15These complexes can be converted to other sulfonic acid salts using conventional techniques (e.g., ionexchange resins, crystallization or ion-pair extraction. These conversion techniques are also useful in purifying the products. Conversion of the pyridine salt to the potassium salt using potassium phosphate or potassium ethyl hexanoate; to the tetrabutylammonium salt using tetrabutyl ammonium hydrogen sulfate; or to a zwitterian (M+ = hydrogen) using formic acid; are particularly useful.

It should be appreciated that the sulfonation reaction which introduces the sulfo group onto the nitrogen atom of the β-lactam nucleus can be effected at various stages of the synthesis, including inser15 tion prior to the formation of a β-lactam nucleus, where such procedure is followed as outlined below.

The sulfonation reaction is effected in the presence of solvents previously described and usually at room temperature. Where the amino function is present it.is preferably conducted with the amino function protected.

Using a benzyloxycarbonyl protecting group as an example, the sulfonation reaction can be visualized as follows: Other protecting groups can be used to protect the amine function, for example, a t-butyloxycarbonyl group, a simple acyl group such as acetyl or benzoyl of phenylacetyl, a triphenylmethyl group, or having the amino function in the form of an azide group. The desired acyl group (R^) can then be affixed by a conventional acylation reaction. 51383 Exemplary acylation techniques for converting a compound of fonnula III to a product of formula I Include reaction with a carboxylic acid (R^-OH), or corresponding carboxylic acid halide or carboxylic acid anhydride. When Rj is alkoxy, acyIcation is best effected by use of an acid chloride or acid bromide.' The reaction with a carboxylic acid proceeds most readily in the presence of a carbodiimide such as dicyclohexylcarbodiimide and a substance capable of forming an active ester in situ such as N-hydroxybenzotriazole. in those instances when the acyl group (R^) contains reactive functionality (such as amino or carboxyl groups) it may be necessary to first protect those functional group* then carry out the acylation reaction, and finally deprotect the resulting product. Alternatively, the sulfonation reaction can be effected with the acyl group already in place, i.e., IV In the case where R2 is lower alkoxy, there can be an additional variation in that the Rj-lower alkoxy group can be inserted after the sulfonation as well as before the sulfonation using the conventional procedure of chlorinating the acylated nitrogen atom in the 3position followed by reaction with a lower alkoxide 513 9 3 -17α. Η acyl-N(Rj) alkoxide Rj «3 aeyl-NB.R-SOj-M*· D-so3~m+ VI The acyl groups in the above reaction also include easily removable groups which function as a protecting group and which can be removed after the reaction to afford the deacylated product (-NHj).

In addition, the β-lactam ring can be formed via a cyclization reaction and the sulfonation reaction can be effected prior to cyclization as well as subsequent thereto, i.e., acyl-NH-kh-so3"m+ cyclisatin VII The acyl group in this reaction can also be an easily removable group which functions as a protecting group and which on removal affords the -NHj product. -18The azetidinone starting materials wherein Rg is hydrogen and at least one of Rg and R* is hydrogen can also be prepared from amino acids having the formula 5 NH2. ty/*4 XII CH-C I C-OH (at least one of Rg and R^ is hydrogen). The amino group is first protected with a classical protecting group, e.g., t-butoxycarbonyl (referred to hereinafter as Boc). The carboxyl group of the protected amino acid is then reacted with an amine salt having the formula + XIII Y-O-NHgCl 2o wherein Y is alkyl or benzyl, in the presence of a earbodiimide to yield a compound having the formula XIV BOC-NH-CH- NH-O-Y 51333 -19(at least one of Rg and R^ is hydrogen). The hydroxyl group of a compound of formula XXV is converted to a leaving group (V) with a classical reagent, e.g., methanesulfonyl chloride (methanesulfonyl is referred to hereinafter as Ms). Other leaving groups (V) which can be utilized are benzenesulfonyl, toluenesulfonyl, chloro, bromo and iodo.

The fully protected compound having the formula BOC-NH-CH15 XV 0' V NH-O-Y (at least one of Rg and R^.is hydrogen) is cyclized by treatment with base, e.g., potassium carbonate. The reaction is preferably carried out in an organic solvent such as acetone, under reflux conditions, and yields a compound having the formula BOC-NHXVI /'3 N-O-Y (at least one of R3 and R^ is hydrogen).

Alternatively, cyclization of a compound of formula XIV can be accomplished without first converting the hydroxyl group to a leaving group. Treatment of a compound of formula XIV with triphenyl phosphine and diethylazodicarboxylate, yields a compound of formula XVI wherein at least one of Rj and R^ is hydrogen. -20Removal of the protecting group from the 1position of an azetidinone of formula XVI can be accomplished via sodium reduction when Y is alkyl, and yields an intermediate having the formula BOC-NH· *4 XVII -NH (at least one of Rj and R^ is hydrogen). If Y is benzyl, catalytic (e.g., palladium on charcoal) hydrogenation will initially yield the corresponding N-hydroxy compound, which upon treatment with titanium trichloride yielda an intermediate of formula XVII wherein at least one of R3 and R4 is hydrogen.

Synthesis involving ring closure of the type described above results in inversion of the stereochemical configuration of the R3 and R4 substituents.

As previously mentioned, the above azetidinone can be sulfonated to fora a compound having the formula XVIII BOC-NH-i-Rj -N-SOj M+ (at least one of Rj and R^ is hydrogen).

An alternative procedure for preparing a compound of formula I wherein Rj is hydrogen and at least one of Rj and R^ is hydrogen, utilizes as a starting material an amino acid amide having the formula nh2-ch0H b |XR4 -C.

XIX -NH. (at least one of Rj and R^ is hydrogen) 21Protection of the amino group with a classical protecting group, e.g., benzyloxycarbonyl (reffered to hereinafter as Z) or Boc, and conversion of the hydroxyl group to a leaving group (V) such as Ms yields a compound having the formula XX ANHOMs -NH, Q*C (at least one or R^ Is hydrogen), wherein A is a protecting group.

Sulfonation of a compound of formula XX yields a compound having the fonnula xxi ANH· (at least one of R^ and Rg is hydrogen).

Cyclization of a compound of formula XXI is accomplished with base, e.g., potassium carbonate. The reaction is preferably carried out in a mixture of water and an organic solvent (e.g., a halogenated hydrocarbon such as 1,2-dichloroethane) under reflux conditions and yields a compound having the formula ΑΝΞXXII n-so3m+ (at least one of and R^ is hydrogen).

Deprotection of a sulfonated azetidinone of formula XXII , wherein A is a protecting group, as well as the counterpart compounds previously described having an R2" alkoxy group by catalytic hydrogenation yields a compound having the formula [2 XXIII N-SO (at least one of Rg and R^ Is hydrogen), wherein R2 is hydrogen or alkoxy which can be converted to the corresponding zwitterion having the formula XXXV nh; -n-so, O'z ., — g (at least one of Rg and R* is hydrogen) by treatment with an acid such as formic acid.

Deprot.ection of a sulfonated azetidinone of formula wherein A is a Boc protecting group using acidic conditions (e.g., using formic acid) yields the corresponding zwitterion of formula An excellent source for the 0-lactam starting materials are the 6-aminopenicillanic acids and the 7-aminocephalosporanic acids which can bear an optional 6 alkoxy or 7-alkoxy substituent respectively. These compounds are of the formulae Ro Rj-NHXXVa XXVb and /CH3 ''•'CHg C-COOH COOH S1393 -23respectively wherein Rg is hydrogen or alkoxy and is hydrogen or acyl. By adapting procedures described in the literature, 3-amino-2-azetidinones can be prepared; see for example, Chem. Soc. Special Publication No. 28, pg. 288(1977); The Chemistry of Penicillins, Princeton Univ. Press, pg. 257, and Synthesis, 494(1977).

The 6-amino-penicillanic acid or 7-aminocephalosporanic acid is first desulfurized by reduction using Raney nickel. The reaction can be run in water under reflux conditions; the resulting compound has the structural formula XXVI Rg-NHN-CH-CH(CH3)2 COOH Replacement of the carboxyl group of the compound of formula XXVI with an acetate group followed by hydrolysis yields a 3-amino-3-alkoxy-2-azetidinone of formula I uherein Rg is hydrogen or acyl, Rg is hydrogen or lower alkoxy, and Rj and R^ are hydrogen. Treatment of a compound of formula XXVI with cupric acetate and lead tetra25 acetate in an organic solvent (e.g., acetonitrile) replaces the carboxyl -group -with-an-acetate group. Hydrolysis of the resulting compound can be accomplished using potassium carbonate in the presence of sodium borohydride.

Introduction of a sulfo group in the 1-position of the above 3-aminp-3-alkoxy-2-e.zetidinone compound can be accomplished by reacting the intermediate with a complex of dimethylformamide and sulfur trioxide. -24A 3-azido-2-azetidinone starting material can be prepared by first reacting an olefin having the formula I* XXVII CH2—C—Rj with a halosulfonylisocyanate (preferably chlorosulfonylisocyanate) having the formula XXVIII O—C—N—SOj-halogen to yield an azetidinone having the formula *4 XXIX N-SOj-halogen Reductive hydrolysis of an azetidinone of formula XXIX yields an N-unsubstituted φ-lactam having the formula *4 XXX Ο*· -NH For a more detailed description of the above described reaction sequence reference can be made to the literature; see, for example, Chem. Soc. Rev., 5, 181 (1976) and J. Org. Chem., 35, 2043 (1970).

An azido group can be introduced in the 3position of an azetidinone of formula XXX (or the sulfonated counterpart) by reaction of the compound with an arylsulfonyl azide (such as toluenesulfonyl azide) to obtain a starting azetidinone having the formula 513 3 3 25R^ XXXI y—HH The reaction proceeds best by first protecting the 5 azetidinone nitrogen with a silyl residue (e.g., t-butyldimethylsilyl, or t-butyldiphenylsilyl), then generating the anion at the 3-position of the nucleus with a strong organic base (e.g., lithium diisopropyl amine) at a low temperature, and then treating the anion with toluenesulfonyl azide. The resulting intermediate is quenched with trimethylsilyl chloride and subsequent acid hydrolysis or fluoride solvolysis of the N-protecting group yields the compound of formula XXXI.

Alternatively, the compound of Formula XXXI can be obtained by first reacting a primary amine having the formula O-alkyl with an aldehyde of the formula R^CH^O to yield the corresponding Schiff base. A[2+2]cycloaddition with an activated form of -azido acetic acid yields a 325 azido-2-azetidinone having the formula R4 •N—Q -26wherein Q is O-alkyl or O-alkyl O-alkyl Oxidative removal of the Q-aubstituent yields the compound of formula XXXI.

The 3-acylamino-2-azetidinones can be obtained by first reducing a 3-azido-2-azetidinone of formula XXXI to obtain the corresponding 3-amino-2-azetidinone and then acylating the 3-amino-2-azetidinone.

Aa previously mentioned in the case where Rj is lower alkoxy, the product can be prepared from the counterpart product wherein Rj is hydrogen.

Clorination of the amide nitrogen of a nonalkoxylated compound yields an intermediate having the formula 51 I 24 XXXII—Λ" -1 [—s Reagents and procedures for N-chlorinating amides are known in the art. Exemplary reagents are tert.-butyl hypochlorite, sodium hypochlorite, and chlorine. The reaction can be run in an organic solvent (e.g., a lower alkanol such as methanol) or in a two phase solvent system (e.g., water/methylene chloride) in the presence of a base such as sodium borate decahydrate.

The reaction is preferably run at a reduced temperature. -27Reaction of an intermediate of formula XXXII with an alkoxylating agent, e.g., an alkali metal alkoxide yields a product of formula I wherein R2 is alkoxy, in combination with its enantiomer. The reaction can be run in an organic solvent, e.g., a polar organic solvent such as dimethylformamide, at a reduced temperature.

An alternative synthesis for preparing the compounds of formula I wherein Rj is alkoxy comprises alkoxylating an intermediate of formula VI wherein R^NH is a carbamate (e.g., Rj is benzyloxycarbonyl) and R2 is hydrogen .

Chlorination of a compound of formula IV using the procedure described above (for chlorination of a non-alkoxylated compound of formula I to yield a compound of formula XXXII yields an intermediate having the formula Cl I XXXIII CgHg-CHj-O-CO-Nj? -N-Cl Using the alkoxylation procedure described above (for converting a compound of formula XXXII to a product of formula I), and subsequently adding a reducing agent such as trimethylphosphite, the compound of formula XXXIII can be converted to an intermediate having the formula )-alkyl R4 XXXIV C^-CRj-O-CO’ NH-NH in a combination with its enantiomer -28The above procedures yield those products of formula I wherein Rj is alkoxy as a racemic mixture. If desired the enantiomer having the R configuration can be isolated from the racemic mixture using conventional techniques such as fractional crystallization of a suitable salt with an optically active organic amine or by ion-paired chromatography utilizing an optically active cation. -29Biologieal Production of the Antibiotic CMS117 Salts of (R)-3-(acetylamino)-3-methoxy-2-oxo»· 1-azetidinesulfonic acid (formula I, R^ is acetyl and Rj is methoxy; referred to hereinafter as EM5117) can also be prepared by cultivation of a strain of the microorganism Chromobacterium violaceum SC 11,378, which has been deposited in the American Type Culture Collection as A.T.C.C. No. 31532.

The Microorganism The microorganism used for the production of EM5117 is a strain of Chromobac ter ium violaceum SC 11,378. A subculture of the microorganism can be obtained from the permanent collection of the American Type Culture Collection, Rockville, Maryland. Its accession number in the repository is A.T.C.C. No. 31532. In addition to the specific microorganism described and characterized herein, it should be understood that mutants of the microorganism (e.g., mutants produced through the use of x-rays, ultraviolet radiation or nitrogen mustards) can also be cultured to produce EM5117.

Chromobacterium violaceum SC 11,378, A.T.C.C.

No. 31532 can be isolated from a moist soil sample containing the microorganism by first stamping 51333 -30the soil sample on a medium containing; Soil Extract - 400 ml Distilled Water- 600 ml 5 Yeast Extract - 5.0 g Glucose -10.0 g Agar -17.5 g The medium is adjusted to pH 6.0 and sterilized 10 in an autoclave at 121°C for 20 minutes. After to 72 hours incubation at 25°C colonies of the Chromobacterium violaceum SC 11,378 are isolated from the plated soil. These isolated colonies are then grown on a medium containing: IS Yeast Extract - 1 g Beef Extract - 1 g NZ amine A - 2 g Glucose - 10 g 20 Agar - 15 g· Distilled water to 1 liter The medium is adjusted to pH 7.3 and autoclaved at 121°C for 30 minutes.

Chromobacterium violaceum SC 11,378 is a gram negative rod often showing barring, bipolar staining and lipid inclusions. It is motile by a single polar flagellum with occassional -31lateral flagella which are smaller in size.

On nutrient agar Chromobacterium violaceum SC 11,378 produces violet colonies. The pigment is enhanced on media rich in tryptophane and yeast extract. In nutrient broth it produces a violet ring on the wall of the tube but no confluent pellicle. The violet pigment is soluble in ethanol but insoluble in water and chloroform. 1° Chromobacterium violaceum SC 11,378 is mesophilic, growing over a range of 15~37°C; no growth occurs at 4 °C or above 37°C. Casein is hydrolyzed strongly by the microorganism, which is oxidase positive. In the presence of Chromobacterium violaceum SC 11,378 glucose, fructose and trehalose are fermented (method of Hugh & Leifson, 1953). L-Arabinose is not utilized by the microorganism either fermentatively or oxidatively. Hydrogen cyanide is produced by 2.0 the microorganism and aesculin hydrolysis is negative.

The above described key characteristics provide the basis for the identification of the microorganism as Chrotnobac ter ium violaceum as distinguished from Chromobacterium lividum, the only other species of the genus Chromobacterium recognized in the 8th edition of Bergey's Manual of Determinative Bacteriology.

Additional strains of Chromobacterium violaceum can also be cultured to produce EM5117. 513 3 3 -32The Antibiotic To obtain the antibiotic EM5117, Chromobacterium violaceuw SC 11,378 A.T.C.C. No. 31532 is grown at, or about, 25°C under submerged aerobic conditions on an aqueous nutrient medium containing an assimilable carbon and nitrogen source.

The fermentation is carried out until substantial antibiotic activity is imparted to the medium, usually about 18 to 24 hours, preferably about hours.

Using the following procedure, EM5117 can be separated from the fermentation medium and purified. After the fermentation has been completed the broth can be centrifuged to remove the mycelium or, alternatively, filtration can be used to remove the mycelium from the broth. After removing the mycelium from the broth EM5117 can be extracted from the broth. Preferably the broth is extracted at pH 5 by ion-pair extraction with cetyldimethyl20 benzylammonium chloride in methylene chloride and back-extracted into aqueous sodium iodide (adjusted to pH 5 with acetic acid). After concentration of the sodium iodide extract in vacuo, an aqueous solution of the residue can be washed with butanol. Concentration of the resulting aqueous solution to dryness yields a residue which is dissolved, to the extent possible, in methanol. Centrifugation can be used to separate insolubles which are washed with methanol and then discarded. -33Purification of the antibiotic can be accomplished by dissolving the methanol concentrate in methanol/water (1:1) and applying it to a chromatography column, e.g., one comprising Sephadex G-10 /Trade Mark) in the same solvent mixture.

After elution with the same mixture active fractions are combined and concentrated to dryness. The residue is mixed with methanol and insoluble material is filtered out and discarded.

Further purification is achieved by applying the methanol soluble material to a column of DEAE cellulose. The column can be eluted with a linear gradient.prepared from pH 5 sodium 0.0. M phosphate buffer and pH 5 sodium 0.1 M phosphate buffer. Active fractions are combined, concentrated and methanol insoluble materials are removed and discarded.

Still further purification of EM5117 is accomplished by dissolving this material in water and placing the solution on a column of Sephadex(Trade Mark)LH-20, and eluting the column with water. Active fractions are combined and con25 centrated. Further purification of EM 5117 is then accomplished by dissolving the material in water, placing the solution on a column of Diaion ΗΡ-20ΑΞ and eluting with water. Active fractions are combined and concentrated. The concentrate is dissolved . in water and passed through a column of Dowex(Trade Mark)50W-X2, potassium form, washing with water.

The effluent can be concentrated to yield a crystalline material 51303 which is relatively pure potassium salt of EM5117.

The above described isolation and purification process yields the potassium salt of EM5117. other salts can be prepared corresponding to the form of the ion exchange resin utilized in the final purification step.

Biological Production of the Antibiotic EM5210 Salts of (R)-3-llN-(D-y-glutamyl)-D-alanyllaminoJ-3-methoxy-2-oxo-l-azetidinesulfonic acid (formula I, is Ο-γ-glutamyl-D-alanyl and Rj is methoxy; referred to hereinafter as EM5210) can also be prepared by cultivation of various acetic acid bacteria.

The Microorganism The microorganism Gluconobacter species SC11,435 can be used for the production of EM5210. A subculture of the microorganism can be obtained from the permanent collection of the American Type Culture Collection, Rockville, Maryland. Its accession number in the repository is A.T.C.C. No. 31581. In addition to the specific microorganism described and characterized herein, it should be understood that mutants of the microorganism (e.g., mutants produced through the use of x-rays, ultraviolet radiation or nitrogen mustards) can also be cultured to produce EM5210. 513S3 -35Giuconobacter species SC11,435, A.T.C.C.

No.31581 can be isolated from ground moss containing the microorganism by first incubating the ground moss in a 10% agueous pectin solution (pH 2.5) for 7 days at 25°C. The organism can then be isolated by plating on a medium containing: •Extract of Spartina patens Grass 400 ml Distilled Water 600 ml Yeast Extract 5 g Glucose 10 g Crude Flake Agar 17.5 g •Extract of Spartina patens grass is prepared by adding 500 g of chopped, dried grass to 3 liters of tap water, brought to a boil and simmered for 30 minutes.

The medium is adjusted to pH 6.0 and sterilized in an autoclave at 121°C for 20 minutes. After about 18 hours incubation at 25°C, colonies of the Giuconobacter species SC11,435 are isolated from the plated pectin enrichment solution. These isolated colonies are then grown on a medium containing: Yeast Extract 1 Beef Extract 1 NZ amine A 2 Glucose 10 Agar 15 Distilled Water to >dium is adjusted to pH at 121 C for 30 minutes.

Giuconobacter'species SCll,435 is a pleiomorphic gram negative rod motile by means -36of one to three polar Flagella. It is obligately aerobic, catalase positive and oxidative. It is differentiated from Pseudomonas in that it is cytochrome oxidase negative and tolerant of extremely acid conditions. These characteristics indicate that the microorganism is more closely related to the acetic acid bacteria than to true Pseudomonads.

On BBL Trypticase soy agar Gluconobacter species SCll,435 grows as a mixture of rough and smooth colony types. The rough type is associated with a faint yellow soluble pigment whereas the smooth type is mucoid and pigmentless. Dissociation between the two types is influenced by media, temperature and storage conditions.

EM5210 activity tends to decrease in cultures where the rough component is dominant.

On BBL Wart agar (pH 4.3) Gluconobacter species SC11,435 grows luxuriantly as heaped up mucoid slimy colonies. Similar growth is-obtained on malt-yeast extract agar (lt each) adjusted to pH 4.5.

On medium containing lt yeast extract, lot glucose, 3t calcium carbonate and 2.5t agar, sufficient acid is produced from the glucose to produce a zone of clearing of the calcium carbonate around the growth. This is a characteristic feature of Acetobacter and Gluconobacter. in the presence of Gluconobacter species SC11,435:(i) a brown soluble pigment is produced on yeast extract-glycerol and yeast extract-calcium -37lactate agar plates, (ii) acid is produced on glucose, fructose, galactose, mannose, xylose, mannitol and arabinose, but no growth or acid production occurs on rhamnose, lactose, sucrose or maltose, when these sugars are incorporated into Hugh and Leifson's medium.

The biological production of Et-15210 is not limited to Gluconobacter species SC11,435, but is broadly distributed throughout the acetic bacteria. The following cultures can each be used for the production of EM5210: Acetobacter pasteurianus subsp.. pasteurianum A.T.C.C. 6033 Acetobacter aceti subsp. aceti A.T.C.C. 15973 15 Gluconobacter oxydans subsp. oxydans A.T.C.C. 19357 Gluconobacter oxydans subsp. suboxydans A.T.C.C. 23773 Gluconobacter oxydans subsp. oxydans 20 A.T.C.C. 15178 Acetobacter aceti subsp. liquefaciens A.T.C.C. 23751 Acetobacter peroxydans' A.T.C.C. 12874 Gluconobacter oxydans subsp. suboxydans 25 A.T.C.C. 19441 Acetobacter sp. A.T.C.C. 21780 Gluconobacter oxydans subsp. industralis A.T.C.C. 11894 -38The Antibiotic To obtain the antibiotic EM5210, Gluconobacter species SC11,435 A.T.C.C. No. 31581 or one of the microorganisms listed above, can be grown at, or about, 25°C under submerged aerobic conditions on an aqueous nutrient medium containing an assimilable carbon and nitrogen source. The fermentation is carried out until substantial antibiotic activity is imparted to the medium, usually about 16 to 24 hours, preferably about 20 hours.

Using the following procedure EM5210 can be separated from the fermentation medium and purified. After the fermentation has been completed the broth is centrifuged to remove the bacteria and the antibiotic is removed from the broth supernateat pH 3.7 by absorption onto an anion exchange resin e.g., Dowex 1-X2. Antibiotic is eluted from the resin with sodium chloride at about pH 4, and the eluate is concentrated and passed through a charcoal column. EM5210 is then eluted from the charcoal with methanol:water,1:1.

The active fractions are collected and dried, and then placed on an anion exchange column, e.g., Dowex 1-X2, and eluted with-a gradient of sodium chloride buffered at pH 4. Active fractions are concentrated and desalted on a macroreticular styrene-divinylbenzene copolymer resin (Dianion HP20AG) column. Active fractions eluted with water are concentrated and freeze dried and this material represents the sodium salt of EM5210. -39The above described purification procedure will vield the sodium salt of EM5210. Other salts can be obtained by changing the salt used to elute EM5210 in the ion-exchange chromatography described above, e.g., using potassium chloride to give the potassium salt.

S1393 -40Example 1 (S)-N-(2-Oxo-l-sulfo-3-azetidinyl)-2-phenylacetamide, potassium «alt Method I: A) 1-I(IR)-carboxy-2-methyl(propyl)]-2-oxo-(3S) [phenyl (acetyl (amino )D azetidine Raney nickel ia washed with water by decantation for several hours until the pH of the water (5-6 times volume of that of the Raney nickel) is 7.6.

To s solution of 9.0 g of penicillin G (Na+) in 500 al of water is added 54 g (90 ml) of Raney nickel. The flask, fitted with a reflux condenser, is immersed in a bath at 155°C. When refluxing begins, it is continued for 15 minutes. The flask is immediately cooled in an ice-water bath, and the Raney nickel is removed by filtration through Celite (Trade Mark.) The pH is adjusted to 3 using dilute HCl, and the aqueous solution is concentrated to ca. 150 ml and cooled. The oily layer crystrallizes upon scratching. After washing with water and drying in vacuo for 3 hours at 50°C there is 3.83 g of the title compound.

B) 1- [Acetyloxy-2-methyl (propyl) ]-2-oxo- (3SHphenyl[acetyl(amino))]azetidine Nitrogen is bubbled for 15 minutes through a stirred suspension of 608 mg (2 mmol) of the above compound in 20 ml of dry acetonitrile.

A water bath at 40-45°C is used for several minutes to dissolve all of the acid. The water 513 9 3 -41bath is removed, and powdered cupric acetate monohydrate (182 mg, 1 mmol) is added, followed after 1 minute of stirring, by 886 mg (2 mmol) of lead tetracetate. The mixture is stirred at roam temperature for 20 minutes. The acetonitrile solution is decanted from the precipitate, and the solids are washed with ethyl acetate. The combined acetonitrile-ethyl acetate solution is evaporated to a residue, which is taken up in ethyl acetate-water. The ethyl acetate layer is washed sequentially with water (3 times), agueous sodium bicarbonate (pH 7), and water.

The ethyl acetate layer is dried over sodium sulfate and evaporated to a residue (515 mg), which is used without further purification in the next reaction.

C) 2-Oxo-(3S)-[phenyl[acetyl(amino)]]azetidine To a solution of 911 mg (2.86 mmol) of the above compound in 21 ml of- methanol is added 3.5 ml of water followed by 383 mg (2.86 mmol) of potassium carbonate. The mixture is stirred under nitrogen for 1 minute, and then 160 mg (4.30 mmol) of sodium borohydride is added. The reaction is stirred at room temperature for 20 minutes. The methanol is removed in vacuo, and the residue is taken up in ethyl acetate and a small amount of water. This is adjusted to pH 2.5. The ethyl acetate layer is washed at pH 7.0 with agueous sodium bicarbonate and then a small volume of water and -42finally dried over eodium sulfate and evaporated to give the crude product (493 mg). Addition of a small amount of ethyl acetate gives 250 mg (43» yield) of the desired crystalline product. Further quantities of product can be obtained by crystallization or chromatography.

D) (S)-N-(2-Oxo-1-sulfo-3-azetidinyl)-2-phenylacetarnide, potassium salt Pyridine-SOj complex (215 mg, 1.35 mmol) is added to a stirred solution of 251 mg (1.23 mmol) of the above product in 2 ml of dry dimethylformamide and 2 ml of dry methylene chloride under nitrogen at rocm temperature. The mixture is stirred for 3 hours. The solvents are removed in vacuo, and the residue is taken up in methylene chloride-water. The pB is adjusted to 6.5 using 2N potassium hydroxide. The aqueous layer is washed vith methylene chloride (3 times) filtered and evaporated to a residue. The residue is stirred with 20 ml of methanol, and the potassium sulfate is removed by filtration.

The filtrate is evaporated to a residue, which is stirred with 10-15 ml of methanol. The solids are collected to give 49 mg of the title compound, melting point 189°C, dec.

Anal. Calc’d for ciiHiiN2°5SKi C, 40.99; H, 3.44; N, 8.69; S, 9.93 Found: C, 45.96; H, 3.83; N, 9.86; S, 8.99 The compound has identical spectral characteristics to the product obtained in Method II.

S 1 39 3 -4 3 Method II; A solution of 660 mg of (S)-2-oxo-3-[I(phenylmethoxy ) carbonyl] amino] -1-azetidinesulfonic acid, potassium salt (see Example 3] in 13 ml of water is stirred in an atmosphere of hydrogen for 2 hours with 200 mg of 10% palladium on charcoal, lhe catalyst is filtered and the filtrate diluted with an equal volume of acetone and cooled in an ice bath. Over 30 minutes, phenylacetyl chloride (eight 40 μ-l portions) and 10% potassium bicarbonate solution are added (pH kept between 5.2-5.8). After 40 minutes, the solution is concentrated in vacuo to remove acetone and applied to a 200 ml HP-20 column. Elution with water and then water-acetone (9:1) gives 160 mg of crude product after TLC examination of Rydon positive fractions, followed by pooling and evaporation. Crystallization from methanol-ether gives 101 mg of the-title compound, melting point 210°C, dec.

Anal. Calc’d for C^Hg^OgSK·1^ HjO: C, 39.86; H, 3.65; N, 8.45; S, 9.68; K, 11.80 Found: C, 40.01; B, 3.37; N, 8.59; S, 9.59; K, 11.98 NMR(D20) 3.66 (s,3), 3.67 (d of fl.., j«6.4), 3.90 (t,j=Kj), 4.90 (d of d, j=6.4) , 7.36 pp® (m.5). -44Method III; To a aolution of (S)-3-amino-2-oxo-lazetidinesulfonic acid, tetrabutylammonium salt (121 mg; aee Example 6A) in dry methylene chloride (3 ml) ia added 40 mg of phenylacetic acid and 61 mg of dicyelohexylearbodiimide. The mixture is stirred for 48 hours at room temperature, and filtered to remove dicyclohexylurea. The solvent is removed in vacuo and the compound residue is taken up in acetone, filtered and the title compound precipitated by the addition of 5 ml of acetone saturated with potassium iodide.

The supernatant is decanted, the residue washed with acetone(3 times) and, after drying, 48 mg of product is obtained, having spectral characteristics in agreement with the products of Methods I and II. Method IV; λ solution of 2.83 g of (3)-2-οχο-3ΓI(phenylmethoxy )carbonyl·]amino]-1-azetidinesulfonic acid, pyridine salt (see Example 2) in 36 ml of water is stirred in an atmosphere of hydrogen with 707.5 mg of 10% palladium on charcoal (175 al of hydrogen taken up). After 2 hours the slurry is filtered, and the filtrate cooled to 0°C and diluted with 46 ml of acetone (initial pH» 4.25 adjusted to pH 6.7 with cold 10% potassium bicarbonate solution). A solution of 2.4 ml of phenylacetyl chloride in 10 ml of acetone is added dropwise over 15 minutes. The pH is maintained between 5.2 to 5.8 by the simultaneous addition of cold 108 potassium bicarbonate solution. After 45 minutes the slurry is 51333 -45diluted with 93 ml of 0.5M potassium phosphate (pH-4.2) and concentrated to remove acetone.

The Blurry is filtered and washed with water.

The filtrate and washings are combined and applied S to a 450 ml HP-20 column. Elution with 1 liter of 0.5M potassium phosphate (pH-4.2), 1 liter of water, and then 2.5 liters of 9:1 water-acetone gives 1.285 g of the title compound in fractions 14-19 (fraction 1-15 were 200 ml, fractions 16-21 were 100 ml). Spectral data is in agreement with that obtained in the foregoing methods. -46Example 2 (S)-2-0XO-3-I[(phenylmethoxy)carbonyl]amino]1-azetidinesulfonic add, pyridine (1;1) salt Method I 1~I(IR)-Carboxy-2-methyl(propyl)]-2-oxo-(3S1[ [ (phenylmethoxy) carbonyl ] amino] azetidine A slurry of 6-aminopenicillanic acid (12.98 g, 0.06 mole) in 140 al of water containing 5.18 g of sodium bicarbonate (stirred for ca. minutes without complete solution) is added in one portion to a well-stirred (mechanical stirrer) suspension of Raney nickel (washed with water to pH 8.0, 260 ml of slurry»130 g) in a 70°C oil bath. After 15 minutes the slurry is cooled, filtered,' and the filtrate treated with 5.18 g of sodium bicarbonate and a solution of 11.94 g (0.07 mole) of benzyl chloroformate in 12 ml of acetone. After 30 minutes, the solution is acidified to pH 2.5 and extracted with methylene chloride. The organic layer is dried, evaporated, and triturated with etherhexane to give a total of 6.83 g of the title compound.

B) 1- [ (Acetyloxy) -2-methyl (propyl) ] -2-oxo-(3S)[ [ (Bhenylmethoxyjcarbonyllaminolazetidine A solution of 6.83 g (0.0213 mole) of the above acid in 213 ml of acetonitrile is treated with 1.95 g (0.0107 mole) of cupric acetate monohydrate and 9.5 g (0.0213 mole) of lead -47tetraacetate. The slurry is immersed in a 65°C oil bath and stirred with a stream of nitrogen bubbling through the slurry until the starting material is consumed. The slurry is filtered and the solids washed with ethyl acetate. The combined filtrate and washings are evaporated in vacuo and the residue taken up in 100 ml each of ethyl acetate and water and adjusted to pH 7. The ethyl acetate layer is separated, dried, and evaporated to give 6.235 g of the title compound.

C) (S)-(2-Oxo-3-azetidinyl)carbamic acid, phenylmethyl ester A solution of 3.12 g (0.0093 mole) of the 15 above acetate in 70 ml of methanol and 7 ml of water is cooled to -15°C and 1.33 g of potassium carbonate and 349 mg of sodium borohydride are added. The reaction mixture is stirred at -15°C - 0°C. After the reaction is complete (ca. 2 hours), the mixture is neutralized to pH 7 with 2N HCl and concentrated in vacuo.

The concentrate is adjusted to pH 5.8, saturated with salt and extracted with ethyl acetate (3 times). The organic layer is dried and evaporated in vacuo. The residue is combined with material from a similar experiment and triturated with ether to give 3.30 g of the title compound. -48D) (S)-2-Oxo-3- ( I (phenylmethoxy) carbonyl lamino] 1-azetidinesulfonic acid, pyridine ealt Method I; A solution of 440 mg (0.002 mole) of the above azetidinone in 2 ml each of dry methylene chloride and dry dimethylformamide is stirred for 2 hours under nitrogen with 35Q mg (0.0022 mole) of pyridine-sulfur trioxide complex. The bulk of the solvent is then removed in vacuo and the residue triturated with ethyl acetate to give 758 mg of solid, which is primarily the title compound.

NMRiDjo-CTsoD) 3.63(1H, d of d, j-6.4), 3.90 (IH, t, j-6), 4.85 (IH, d of d, ;»6.4), 5.10(22,S) 7.27 (5H,S) 8-0-9.oppmfm’s 5H).

Method Hi Chlorosulfonyltrimethylsilyl ester (18.87 g) is added dropwise at -20°C to 7.9 g of anhydrous pyridine with stirring under a nitrogen atmosphere. When the addition is complete, stirring is continued for 30 minutes at room temperature, and trimethylchlorosilane is then removed in vacuo, λ solution of 20 g of the above azetidinone (Method I, part C) in 120 ml of dimethylformamide and 120 al of methylene chloride is added and stirring at ambient temperature is continued for θ 3.5 hours. The solvent is distilled off in vacuo and the oily residue crystallized by addition of ethyl acetate, yielding 31 g of the title canpound. nmr data is identical to that of the Method I product. 51333 -49Example 3 (S)-2-Oxo-3-[I(phenyImethoxy)carbonyl]amino]l-azetidinesulfonie acid, potassium salt Method I: (S)-2-OXO-3-II(phenylmethoxy)carbonyl] amino]1-azetidinesulfonic acid, pyridine salt (135 mg; see Example 2) is dissolved in 2 ml of 0.5 M monobasic potassium phosphate (adjusted to pH 5.5 with 2N potassium hydroxide) and applied to a 25 ml HP-20AG column. The column is eluted with 100 ml of buffer, 200 ml of water and 100 ml of 1:1 acetone-water. Fractions (25 ml) 14-15 are highly Rydon positive. Evaporation yields 80 mg of material which is primarily the title compound(spectral data identical to that prepared below) Method II: (S) -2-0XO-3- [ [ (phenylmethoxy) carbonyl] amino] 20 1-azetidinesulfonic acid, pyridine salt (600 mg; see Example 2) is dissolved in 2 ml of water and mixed with 15 ml of pH 5.5 monobasic potassium phosphate buffer. A solid forms and the* slurry is cooled to 0°C, filtered, washed with cold buffer, cold 50% ethanol, ethanol and ether to give 370 mg of the title compound (containing excess potassium ion by analysis). A solution of 280 mg of the salt in 10 ml of water is applied to a 100 ml ΞΡ-20 column. The column is eluted with 200 ml of water and then water-acetone (9:1). -50Fraetiona (50 ml) ara collected; evaporation ot fraction 7 give· a solid. Trituration with acetone, filtration, and drying in vacuo gives 164 mg of the title compound, melting point 193-196°C.

Anal. Calc’d for ciihhN2O6SK’1/2H2O; C, 38.02; B, 3.48; N, 8.06; S, 9.23; X, 11.25 Found: C, 38.19; B, 3.24; N, 8.15; S, 9.12; X, 11.53 NMR(D20) 3.69(1H, d, of d, ;-6.4), 3.91(1H, t, ;»6) , 4.76(18, m), 5.16(2H,S), 7.43ppm (5h,S) Method III: (5)-(2-Oxo-3-azetidinyl)carbamic acid, phenylmethyl ester (20.0 g, see Example 2C) is suspended in 200 ml of acetonitrile, 21.6 ml of monotrimethylsilyltrifluoroacetamide (25.3 g). is added and the mixture is heated to 50°C with stirring for 1 hour. After cooling in an ice bath to 0°C, 17.2 g of trimethylsilyl ehlorosulfonate is dropped in and the solution is stirred at ambient tenperature for 6 hours. To the solution is added 24.2 g of potassium ethyl hexanoate in 100 ml of butanol and stirring is continued for en additional 1 hour. The slurry is poured into 1 liter of dry diethyl ether and the precipitate ia filtered off and dried in vacuo. The compound is dissolved in 500 ml of water, the pB is adjusted to 5.0 with potassium carbonate, insoluble material is filtered off and the mother liquor is freeze dried. The yield of crude canpound is 19.4 g. The compound contains small amounts of potassium chloride which is removed by chromatography,spectral data identical to that of Method II. 5139 3 -51Zxample 4 (S)-2-Oxo-3-I[(phenylmethoxy)carbonyl]amino]1-azetidinesulfonic acid, tetrabutylammonniura salt (1:1) Method I; (S)-2-QXO-3-[I(phenylmethoxy]carbonyl] amino] 1-azetidinesulfonic acid, pyridine salt (1:1) (34.3 g; see Example 2) is dissolved in 800 ml of water. The solution is cleared with charcoal, .7 g of tetrabutylammonium hydrogen sulfate in 80 ml wafer is added and the pH is adjusted to .5 with iN-potassium hydroxide. The solvent is removed in vacuo until a volume of about 200 ml X3 is reached. The precipitated tetrabutylammonium salt is filtered off and dried in vacuo. The compound can be recrystallized from water or dissolved in methylene chloride, filtered and precipitated by addition of ether. Yield 34.3 g melting point 108-110°C.

Method XI: (S)-2-QXO-3-iI(phenylmethoxy)carbonyl] amino] 1-azetidinesulfonic acid, potassium salt (1:1) (20.2 g; see Example 3) is dissolved in 500 ml of water, filtered and 20.3 g of tetrabutylammonium hydrogen sulfate in 100 ml of water is added. The pH is brought to 5.5 with IN potassium hydroxide. The volume is reduced in vacuo to about 100 ml and the precipitated tetrabutylaromonium salt is filtered off. The compound is -52S1393 dissolved in 30 ml of methylene chloride, filtered and precipitated by addition of ether, yielding 21 g of the title compound, melting point 109-lll°C.

Example 5 (3S)-a-I[(2-Oxo-l-sulfo-3-azetidinyl)amino]carbonyl]benzeneacetic acid, phenylmethyl ester, potassium «alt (1:1) λ) (S)-3-Amino-2-Azetidlnone (S)-(2-Qxo-3-azetidinyl)carbamic acid, phenylmethyl ester (3 g; see Example 2C) is hydrogentated in 100 ml of methanol in the presence of 1 g of palladium on charcoal catalyst. When the theoretical amount of hydrogen is absorbed, the catalyst is filtered off and the filtrate evaporated to dryness. On standing, 1.1 g. ot the title compound crystallizes.

B, (3S)-a-[I(2-Oxo-3-azetidinyl)amino]carbonyl]benzeneacetic acid, phenylmethyl ester The above azetidinone (3.0 g) is dissolved in 100 ml of dimethylformamide. The solution is cooled to 0°C and 4.5 g of N-methylmorpholine is added followed (dropwise) by 10.8 g of a-(chlorocarbonyl)benzeneacetic acid, phenylmethyl ester in 50 ml of acetonitrile with stirring. The mixture is stirred for about 16 hours at 5°C. The solvent is distilled off in vacuo and 100 ml of water is added to the residue. The agueous suspension is extracted twice with 100 ml portions of methylene chloride. -53The organic layers ore combined, washed with sodium bicarbonate, 2N phosphoric acid and water, dried with eodium sulfate, filtered and evaporated to dryness. The residue is crystallized with ethyl acetate and petroleum ether, yielding 8.7 g, melting point 164-166°C.

C) (3S)-a° [ f (2-Qxo-l-sulfo-3-azetidinyl)an)inojcarbonyl]benzeneacetic acid, phenylmethyl ester, potassium salt (1:1) The above compound (8.9 g) is suspended in 150 ml of acetonitrile. Monotrinethylsilyltrifluoroacetamide (5.7 g)is added and the solution is heated for 30 minutes at 50°C with stirring. The solution is cooled to 0 C and 3.9 g trimethylsilyl chlorosulfonate is added dropwise. When the addition is complete the mixture is heated to 50°C for 5 hours.

After cooling to 20°C, 7.δ g of potassium ethyl hexanoate in 10 ml of butanol is added and stirring is continued for 30 minutes. On addition of 300 al of ether the title compound precipitates and is filtered off. The crude product is stirred with 100 ml of dry acetonitrile for 30 minutes and filtered off, yielding 4.5 g of the title compound, melting point 118-120°C. Further purification of the crude product by HP20 chromatography followed by freeze-drying yields pure material having a meltinc point of 188-190’C. -54Exaraple 6 (5)-3-1((2-Amino-4-thiazolyl)acetyl]amino]-2oxo-l-azetidinesulfonic acid, potassium ealt A) (S)-3-Aiaino-2-oxo-l-azetidinesulfonic acid, tetrabutylammonium salt (5, -2-0xo-3-11 (phenylmethoxy) carbonyl) aminoj 1- azetidinesulfonic acid, tetrabutylammonium salt (2 g; see Example 4) is dissolved in 100 ml of dimethylformamide and hydrogenated for about 30 minutes with 1 g of palladium on charcoal (10») as catalyst. The catalyst is filtered off and the dimethylformamide is removed leaving the title compound as an oil. NMR (CDClj) 3.82 (IB, t,j« 5.5), 4.05(d. IB, d of d, )· 5.5, 2.5 cps,.

B) (S)-3-[[(2-Aaino-4-thiazolyl)acetyl]aaino]2- oxo-l-azetidinesulfonic acid, potassium salt The above compound (2 g), 0.5 g of aminothiazole acetic acid and 0.4 g of hydroxybenzotriazole are stirred at 0°C in 100 ml of dry dimethylformamide, while a solution of 0.7 g of dicyclohexylcarbodiimide in 10 ml of dimethylformamide is added dropwise. After the addition is complete, stirring is continued for 12 hours at 20°C. Insoluble urea is filtered off and the solvent is evaporated in vacuo. The oily residue is treated with a solution of potassium perfluorobutane sulfonate in 20 ml of acetone at room temperature for 15 minutes. After the addition of 200 ml of dimethyl ether the title compound precipitates, and is filtered off, dried and purified via a 300 ml BP-20 chromatography column using water as eluent. The yield 51333 -55is 850 mg of the title compound, melting point >300°C.

Example 7 [3S (1))-3-(((FormyloxyIphenylacetyl]amino]-2oxo-l-azetidinesulfonie acid, potassium salt (S)-3^Amino-2-oxo-l-azetidinesulfonic acid, tetrahutylammonium salt (1.5 g; see Example SA) in 100 ml of dimethylformamide and 2 ml of propylene oxide are cooled to 0°C.

A solution of O-formylmandelic acid chloride in 10 ml of acetonitrile is added dropwise with stirring. The temperature is maintained for 1 hour and the solvent is then distilled off IS in vacuo. The oily residue is treated with a solution of 2 g of potassium perfluorobutane sulfonate in 15 ml of acetone. After adding 200 ml of ether, the title compound crystallizes and is filtered off yielding 1.5 g of product. The product is purified by HP-20 chromatography, m.p. 180-185°C. with decomposition.

Example 8 [33(+)]-3-11 (Formyloxy)phenylacetyl]amino]2-oxo-l-azetidlnesulfonic acid, potassium salt 2S Following the procedure of Example 7, but substituting D-O-formylmandelie acid chloride for O-formylmandelic acid chloride, yields the title compound, melting point 120-125°C, after freeze drying. -56513S3 Example 9 [3S (-))-3- [ I (Forayloxy)phenylacetyl] amino-2oxo-l-azetidinesulfonic acid, potassium salt Following the procedure of Example 7, but substituting L-O-formylmandelic acid chloride for O-formylmandelie acid chloride, yield· the title compound, containing 1 mole of water, melting point, 203-205°C. After careful drying the product melt· at 228-230°C.

Example 10 (S) -2-ΟΧΟ-3-1 (1-oxooctyl) amino] -1-azetldinesulfonic acid, potassium salt (S) -3-Amino-2-oxo-l-azetidinesulfonic acid, tetrabutylammoniun salt (1.5 g; see Example 6A) in 100 ml of dimethylformamide and 2 ml of propylene oxide are cooled to 0°C. At this temperature a solution of 0.8 g of caprylic acid chloride in 20 ml of dry acetone is added dropwise and stirring is continued for 30 minutes. The solvent is evaporated in vacuo, and the oily residue is treated vith 2 g of potassium perfluorobutane sulfonate in 15 ml of acetone. The acetone is distilled off in vacuo. The residue is dissolved in 5 ml of water and chromatographed using 300 ml of HP-20 resin and water/acetone (9:1) as eluent, yielding 0.9 g of the title compound, melting point 173-180°C, after freezedrying . -57Example 11 I3S(Z)]-3-[[(2-aroino-4-thiazolyl)([(hydroxy(phenylmethoxy) phosphinyl jmethoxy]imino]acetyl]amino]2-oxo-l-azetidinesulfonic acid, potassium salt (S)-3-toino-2-oxo-l-azetidinesulfonic acid, tetrabutylammonium salt (0.8 g; see Example 6a) in 30 ml of dimethylformamide, 0.9 g of (2)-2amino-s-[I[hydroxy(phenylmethoxy)phosphinylJmethoxy]imino]-4-thiazoleacetic acid, 0.3 g of hydroxybenzotriazole and 0.7 g of dicyclohexylcarbodiimide are stirred for 24 hours at room temperature. The precipitated urea is filtered off and the solvent is removed in vacuo. The reamining oil is treated with an equivalent amount IS of potassium perfluorobutane sulfonate in 10 ml of acetone. The title compound is filtered off and purified using HP-20 resin and water as eluent, yielding 500 mg, melting point 210-215°C,dec Example 12 [3S(Z)]-3-[[(2-Amino-4-thiazolyl)(ethoxyimino)acetyl]amino]-2-oxo-l-azetidinesulfonic acid, potassium salt ' (S)—3—Amino—2—oxo—1—azetidinesulfonic acid tetrabutylammonium salt (1.5 g; see Example 6A) in 100 ml of dimethylformamide, 0.6 g of hydroxybenzotriazole , 1 g of dicyclohexylcarbodiimide and 0.8 g of (Z)-2-amino-a-(ethoxyimino)-4thiazoleacetic acid are stirred at roam temperature for 24 hours. The solvent is distilled off and the -residue is dissolved in 30 ml of acetone. -58Urea is filtered off and the mother liquor is treated with a solution of 2 g of potassium perfluorobutane sulfonate in 20 ml of acetone. After the addition of 200 al of ether the title compound precipitates, is filtered off and dried. Purification is accomplished by chromatography using an BP-20 column and water as eluent, yielding 1.1 g of thg title compound, melting point 180-185°C, dec.

Example 13 [3S(B)]-3-II(2-Amino-4-thiazolyl)(ethoxyimino)acetyl]amino]-2-oxo-l-azetidinesulfonic acid, potassium salt Following the procedure of Example 12, but substituting (E)-2-amino-a-(ethoxyimino)4-thiazoleacetie acid for (Z)-2-amino-a-(ethoxyimino) -4-thiazoleacetic acid, yields the title compound, melting point 160-170°C after freeze drying.

Example 14 (3S(Z)-3-I[(2-Amino-4-thiazolyl)](2,2,2-trifluoroethoxy) imino]acetyl]amino]-2-oxo-l-azetidinesulfonic acid, potassium salt Following the procedure of Example 12, but substituting (Z)-2-amino-a-[(2,2,2-trifluoroethoxy)imino]-4-thiazoleacetic acid for (Z)-2-amino«-(ethoxyimino)-4-thiazoleacetic acid, yields the title compound melting point 160-170°C after freeze-drying.

Example IS (S)-2-0XO-3-((1-oxopropyllamino]-1-azetidinesulfonic acid, potassium salt Method I: (S)-3-Asjino-2oxo"l"azetidinesulfonic acid, tetrabutylammoniisn salt (1.5 g; see Example SA) in 100 ml of dry dimethylformamide and 4 ml of propylene oxide are cooled to 0°C with stirring.

At this temperature 0.5 g of propionic acid chloride in 10 ml of acetonitrile is added dropwise and the mixture is stirred for 2 hours.

The solvent is distilled off in vacuo and the oily residue is treated with an equivalent amount of potassium perfluorobutane sulfonate in 5 ml of acetone. On addition of ether the title compound crystallizes and is filtered off, yielding 0.8 g of product, melting point 135-140°C after freeze-drying.

Method II: (S) -2-CKO-3- ϊ I (phenylmethoxy) carbonyl] amino] 1-azetidinesulfonic acid, tetrabutylammonium salt (4 g; see Example 4) is hydrogenated in 100 ml of diglyme with 1 g of palladium on charcoal.

The hydrogenation is complete after 2.5 hours.

The catalyst is filtered off and 2 ml’ of propylene oxide is added. After cooling to 0°C,0.5 g propionic acid chloride in 10 ml dry diglyme is added with stirring. After 30 minutes the solvent is removed in vacuo and the oily residue is treated -60with an equivalent amount of potassium perfluorobutane sulfonate in 20 ml of acetone. After the addition of ether, the title compound crystallizes, is filtered off and recrystallized from water/acetone, yielding 0.9 g of product, melting point 156-160°C (dec.).

Example 16 [3S(±)3—3—I (Hydroxyphenylacetyl)amino]-2-oxo1-azetidinesulfonic acid, potassium salt (S)-3-Amino-2-oxo-l-azstidine>ulfonic acid, tetrabutylammonium salt (1.5 g; see Example 6A) in 100 ml of dry dimethylformamide is stirred for about 16 hours with 1.5 g of dicyelohexylearbodiimide, 0.5 g of hydroxybenzotriazole and 0.6 g of mandelic acid. The solvent is removed in vacuo and the residue is dissolved in 20 ml of acetone. The precipitated urea is filtered off and the mother liquor is treated with an equivalent amount of potassium perfluorobutane sulfonate. After the addition of ether the title compound precipitates and is filtered off, yielding 1.4 g of crude product.

After recrystallization from water, the product has a melting point of 138-140°C.

Example 17 (S)—3— f[[(Cyanomethyl) thio]acetyl] amino]-2-oxol-azetidinesulfonic acid, potassium salt (S)-3-Araino-2-oxo-l-azetidinesulfonic acid, tetrabutylammonium salt (1.5 g; see Example 6A) and 0.72 g of [(cyanomethyl)thio]acetic acid are dissolved in 70 ml of acetonitrile and a solution of 1.04 grams of dicyelohexylearbodiimide -61in acetonitrile is added dropwise. The mixture is stirred for about IS hours at 0°C and the precipitated dicyclohexylurea is filtered off, the filtrate evaporated and the oily residue dissolved in acetone. On the addition of a saturated solution of potassium iodide in acetone, the title compound precipitates, yielding 1.1 g of product, melting point 150-155°C.

Example 18 (S)-2-Oxo-3-[ (lB-tetrazol-1-yl ace tyl) amino 1^-1azetidinesulfonie aeid, potassium salt To a solution of 0.005 mole of (S)-3-amino2-oxo-l-azetidinesulfonie acid, tetrabutylammonium salt (see Example 6A, in 70 ml of dimethylformamide is added 0.77 g of IH-tetrazole-l-acetic acid and a solution of 1.13 g of dicyclohexylcarbodiimide in 5 ml of dimethylformamide. The mixture is stirred for about 16 hours at room temperature and the solvent is removed in vacuo. The remaining oil is dissolved in 20 ml of acetone and treated with 0.006 mole of a solution of potassium perfluorobutane sulfonate in acetone, yielding 1.5 g of the title compound, melting point 170-175°C, dee.

Example 19 (S)-2°Oxo-3- [ (2H-tetrazol-2-ylacetyllamino]rlazetidinesulfonic acid, potassium salt Following the procedure of Example 18, but substituting 2H-tetrazole-2-acetic acid for IH-tetrazole-l-acetic aeid, yields the title compound, melting point 175-177 C, dee. -62Exawple 20 (S)-2-OXO-3-I(2-thienylacetyliaainol-l-azetidinesulfonic acid, potassium salt Following the procedure of Example 18, but substituting 2-thiopheneacetic acid for lE-tetrazole-l-acetic acid, yields the title compound, melting point 180-190°C, dec.

Example 21 I3S(Z)]-3-[[(2-Amino-4-thiazolyl) (methoxyimino)acetyl] amino] -2-oxo-l-azetidinesulfonic acid, potassium salt (S)-3-Araino-2-oxo-l-azetidinesulfonic acid, tetrabutylammonium salt (prepared as described in Example 6A from 7.9 g of (S)-2-oxo3- [ I (phenylmethoxy)carbonyl] amino] -1-azetidinesulfonic acid, tetrabutylammonium salt) is cooled to 0°C and 3.53 g of (Z)-2-amino-β-(methoxyimino)4- thiazoleacetic acid is added, followed by a solution of 3.27 g of dicyclahexylcaxbodiimide in 10 ml of dimethylformamide. The mixture is stirred for 16 hours at 5°C, filtered and the solvent removed in vacuo. The residua is dissolved in acetone and filtered. On addition of 60 ml of a 10% solution of potassium perfluorobutane sulfonate in acetone, 4.7 g of crude product crystallizes. The crude product is purified by chromatography on HP-20, 100-200 mesh to yield 3.0 g of the title compound, melting point 235°C. -63Example 22 <3S)—c — 11(2-Oxo-l-Bulfo-3-azetidinyl)amino]carbonyl]benzeneacetic acid, dipotassium salt (3S)-e-[I(2-Oxo-l-sulfo-3-azetidinyl)amino]5 carbonyl]benzeneacetic acid, phenylmethyl ester, potaesium salt (100 mg; see Example 5) is dissolved in 20 si of anhydrous methanol. Palladium on charcoal (10 mg, 10%) is added and the mixture is treated with hydrogen for 15 minutes. The 1® catalyst is filtered off, and the methanol evaporated in vacuo. The residue is dissolved in 5 ml of water and the pH adjusted to 6 with IN potassium hydroxide. After freeze drying crude product is obtained. The crude material I5 is chromatographed over HP 20 resin (water as eluent), yielding 60 mg of the title compound melting point 80-85°C.

Example 23 (S)-3-(Acetylamino)-2-oxo-l-azetidinesulfonic acid, potassium salt Method I: A) (S)-3-Amino-2-oxo-l-azetidinesulfonie acid, potassium salt (S)-3~(Benzyloxycarbonylamino)-2-oxo-l25 azetidinesulfonic acid, potassium salt (169 mg; see Example 3) is dissolved in 4.0 ml water and hydrogenated over 37 mg of 10% palladium on charcoal for 1 hour 40 minutes. The catalyst is removed by filtration and washed with 1 ml of 3® 50% aqueous acetone. -64B) (S)-3-(Acetylamino)-2-oxo-l-azetidinesulfonic acid, potassium salt The above solution of the free amine is diluted with 3.5 ml of acetone and stirred in an ice bath. Acetyl chloride (320 pi} is added over ca. 15 minutee in small portions and alternated with solid potassium bicarbonate to maintain pH 6.5-7.2. After 30 minutes e silica gel TLC in acetone:.acetic acid (19:1) and the Bydon test indicated the reaction to be essentially finished. Six ml of 0.5M pH 5.5 monobasic potassium phosphate buffer are added and the solution is acidified to pH 4.8 with 2N hydrochloric acid. Tbe acetone is removed in vacuo and the aqueous solution thus obtained is passed through a 50 ml HP-20 AG column to give 2.197 g of solid. This is digested with methanol to yield 282 mg of extractable material which still containssome salt. The product is further purified by passage through an ISC-50 column, followed by acidification to pH 3.8 with subsequent stripping to dryness in vacuo. Trituration with acetone gives 64 mg of desired product containing ca. 0.5 equivalent of inorganic potassium salts. Final passage through a 200 ml HP-20 AG column, followed by lyophilization from .5 ml of water gives 22 mg of product as amorphous powder which is dried in vacuo for 2 hours at 50°C, melting point 170-180°C after softening at 100°C.

Anal, for CjH^NjSK, Calc*d: C, 24.38; H, 2.87; N, 11.37; X, 15.9 Found; C, 26.06; H, 3.14; N, 9.96; K, 18.04 65Mefchod 21: A solution of 2.0 g of (S)-2-oxo-3-II(phenyl methoxylcarbonyl]amino]-1-azetidineuslfonic acid, pyridine salt (see Example 2) in 25 ml of water § ie hydrogenated over 500 mg of 10% palladium on charcoal. After 2 hours the solution is filtered, cooled to 0°C and 40 ml of acetone is added.

The pH of the solution is kept between 5.2-5.8 by simultaneous addition of acetyl chloride and cold % potassium bicarbonate solution. The pH of the solution is adjusted to 4.2 with acetyl chloride and the solution is concentrated on the rotary evaporater to remove acetone. Chromatography on a 300 ml BP-20 AG column (water eluant -25 ml fractions) gives 900 mg of the title compound in fractions 13 and 14 contaminated with some potassium acetate. Rechroraatography on EP-20 AG gives an analytical sample, melting point 205-210°C.

Anal.calc'd for CgH^OjSK: C,24.38, B,2.86; N.11.38; 3,13.02; K,15.88 Found; C.24.23; B.2.81; »,11.25; S,12.86; K.15.74 513 9 3 Example 24 (S)-2-Oxo-3-f(phenoxyaeetyllamino]-1-azetidinesulfonic acid, potassium «alt (S)-3-Amino-2-oxo-l-azetidinesulfonic acid, tetrabutylammonium salt (1.5 g; aee Example 6A) in 100 ml of dry dimethylformamide ie cooled to 0°C. Propylene oxide (2 ml) is added and 1 g of phenoxyaeetyl chloride ie added dropwise with stirring. The reaction is completed within 1 hour. The solvent is removed in vacuo and the residue ie treated with as equivalent amount of potassium perfluorobutane sulfonate in 20 ml of acetone. On addition of ether the title compound (1 g) crystallizes and is filtered off and dried. After recrystallization from boiling water the title compound has a melting point of 176-180°C.

Example 25 (3S(R*))-3-(([[[3-[(2-Furanylmethylene)amino]-2oxo-l-imidazolidinyl]carbonyl]amino)phenylacetyl]amino]-2-oxo-l-azetidinesulfonic acid, potassium salt (S) -2-0XO-3-1 [ (phenylmethoxy) carbonyl] amino]1-azetidinesulfonic acid, tetrabutylammonium salt (3 g; tee Example 4) is hydrogenated in 100 ml of dimethylformamide with 1.5 g of palladium on charcoal. After 0.5 hours, the catalyst is filtered off and 1.8 g of dicyclohexylcarbodiimide, 2 g of (R)-α-(I[3-1 (2-furany1methylene) amino] -2-oxo-l-imidazolidinyl] carbonyl]51393 -67amino]benzeneacetic ocid and 0.9 g of hydroxybenzotriazole are added. After 3 hours the reaction mixture is evaporated to dryness, dissolved in 50 ml of dry acetone and the § precipitated urea removed by filtration. An equivalent amount of potassium perfluorobutane sulfonate in 20 ml of acetone-is added and the title compound precipitates. Crystallization is completed by addition of 200 ml ether.

After filtration the title compound is recrystallized from water, yielding 2 g, melting point 220-225°C, dec.

Example 26 IS [35(S*)]-2-Oxo-3-lift(2-oxo-l-imidazolidinyl) carbonyl) amino]phenylacetyllamino]-1-azetidinesulfonie acid, potassium salt (S) —2—Oxo—*3—[ [ (phenylmethoxy)carbonyljamino]1-azetidinesulfonic acid, tetrabutylammonium salt (3 g; see Example 4) is hydrogenated in 100 ml of dry dimethylformamide with 1.5 g of palladium on charcoal; the catalyst is filtered ©ff after 30 minutes. (E)-s-[[(2-Oxo-l-imidazolidinyl) carbonyl] amino] benzeneacetic acid (1.8 g), 1.3 g of dicyclohexylcarbodiimide and 0.9 g hydroxybenzotriazole are added and the solution is stirred for 2.5 hours. The solvent is removed in vacuo and theresidue is dissolved in 50 ml of acetone. The precipitated urea is filtered off and the mother liquor is treated with an equivalent -68•mount of potassium perfluorobutane sulfonate in 20 ml of acetone. The title compound crystallizes and ia filtered off after the addition of 200 ml of ether, yielding 1.8 g of product, melting point 210-215°C after recrystailization from water/acetone.

Example 27 135(2)3—3—11(Methoxyimino)phenylacetyl]amino]-2oxo-l-azetidlnesulfonic acid, potassium salt A) [3S(2)]-3- (I (Methoxyimino)phenylacetyl] amino]2-azetldinone (Z)-a-(Methoxyimino)benzeneacetic acid (3.58 g) ia dissolved in methylene chloride, cooled to 5°C, end treated with e solution of 4.53 g of dicyclohexylcarbodiimide in 50 ml of methylene chloride. The mixture is stirred for 30 minutes et 5°C end a solution of 1.72 g of 3-emino-2-azetidinone (see Example 5A) in 100 ml of methylene chloride is added. The reaction mixture is kept at 5°C for 1 hour and for 2 hours et rocm temperature. After removal of the dicyclohexylurea by filtration, the filtrate is evaporated, yielding 6.6 grams of crude product. This material is purified by chromatography on 750 grams of silica gel, using a mixture of methylene chloride/ethyl acetate (7:3) as eluent, yielding 2.9 g of product.

S1393 -69B) (3S(Ζ))3 —3—(Γ(Methoxyimino)phenylacetyl]aminoJ-2-oxo-l-azetidinesulfonic acid, potassium salt Pyridine (0.5 ml) is dissolved in 5 ml of 5 absolute methylene chloride, cooled to -30° C and a solution of 0.93 ml of trimethylsilyl chlorosulfonate in 5 ml of methylene chloride is added. The mixture is stirred at room temperature for 30 minutes and evaporated in vacuo to dryness. The residue is dissolved in 10 ml of dimethylformamide and treated at room temperature with a solution'of .1.23 g of the above azetidinone in 10 ml of dimethylformamide. After stirring for two hours at room temperature, the solution is evaporated to dryness to yield 2.1 g of crude [3S (Z))-3-( [ (methoxyimino) phenylacetyl] amino] 2-oxo-l-azetidinesulfonic acid, pyridine salt.

Treatment of tbe pyridine salt with tetrabutylammonium hydrogen sulfate yields the corresponding tetrabutylammonium salt which is extracted with methylene chloride and remains as an oil after evaporation.

Treatment of the tetrabutylammonium salt with an equimolar amount of potassium 2S perfluorobutane sulfonate in acetone, evaporation, and treatment of the residue with ether, yields 1.6 grams of the title potassium salt which is purified by chromatography on HP-20. Elution is carried out with water/acetone 90:10 and yields a product having a melting point of 220°C,dec. -70Example 28 [3S(Z)]-3-If(2-Amino-4-thiazolyl)f f2-(diphenylmethoxy)-l,l-dimethyl-2-oxoethoxy]imino]acetyl]amino]-2-oxo-l-azetidinesulfonie acid, potassium salt (1.-1) λ solution of 0.005 mole of (S)-3-amino2-oxo-l-azetidinesulfonic acid, tetrabutylammonium salt (see Example 6a) and 0.006 mole of (Z) -2-amino-o-112- (dipheny lmethoxy) -1, 1-dimethyl1® 2-oxoethoxy] imino]-4-thiazoleacetic acid in 60 ml of dimethylformamide is treated with 0.7 g of hydroxybenzotriazole and 1.13 g of dicyclohexylcarbodiimide. The mixture is stirred for about 16 hours at room temperature, filtered and the filtrate evaporated. The residue is dissolved in 30 ml of acetone, filtered and treated with 20 ml of a solution of 10% potassium perfluorobutane sulfonate in acetone. After the addition of petroleum ether the title compound precipitates 2® and is treated with ether and filtered to yield 3.8 g of product,melting point 190°C, dec.

Example 29 [3S(Z)1-3-(1(2-Amino-4-thiazolyl)[(1-carboxy-l25 methylethoxy) imino]acetyl]amino)-2-oxo-l-azetidinesulfonic acid, dipotassium salt (3S(Z)]-3-1[(2-Amino-4-thiazolyl) [ 12(diphenylmethoxy)-1,1-dimethyl-2-oxoethoxyJ imino]acetyllamino]-2-oxo-l-azetidinesulfonic acid, potassium salt (2 g; see Example 28) is suspended in 5 ml of anisole and 25 ml of trifluoroacetic — acid is added at -10°C. The mixture is stirred for 10 minutes at -10^C. Ether (100 ml) is added slowly at -10°C and subsequently 50 ml of petroleum ether is added. The precipitate is filtered to' yield 1.6 g of the trifluoroacetic acid salt. This is suspended in 20 ml of water at 0°C, adjusted to pH 5.5 with diluted potassium hydroxide and purified on an HP-20 column. The title compound is eluted with water and has a melting point of 225°C,dec.

Example 30 I3S(2)]-3°I[(2-Aroino-4-thiazolyl)[{2-(diphenylmethoxy) -2-oxoethoxy Umino] acetyl] amino] -2-oxol-azetidinesulfonic acid, potassium salt Following the procedure 'described in Example 28, but substituting (Z)-2-amino-a-([2(diphenylmethoxy)-2-oxoethoxy] imino] -4--thiazoleace tic acid for (Z)-2-amino-a-[ [2-(diphenylmethoxy) 1, l-dimethyl-2-oxoethoxy] imino] -4-thiazoleacetic acid, yields the title compound, melting point 180°C. dec. -72Examplc 31 [3S(±)3~3I (Azidophenylacetyl) amino]-2-oxo-lazetidinesulfonic aeid, potassium salt Method 1: A) (i,S)-a-Azido-N-(2-oxo-3-azetldinyl)benzeneacetamide (S)-3-Amino-2-azetidinone (2.15 g; see Example 5A) and 2.1 g of sodium bicarbonate q axe dissolved in 50 ml of acetone/water (2:1). (±)-a-Azidobenzeneacetyl chloride (5 g) dissolved in 10 ml of acetone is added dropwise while maintaining the temperature at 0-5°C and the pH at 6.B with sodium bicarbonate. After stirring for 1 hour, the acetone is distilled off and the remaining aqueous solution is adjusted to pH 8 with sodium carbonate and extracted with three 50 ml portions of methylene chloride. Evaporation of the sodium sulfate dried organic layers yields 3.6 g of the title compound as an oil which crystallizes after triturating with ether. Recrystailization from methylene chloride/ether, yields the title compound, melting point 97-100°C. 13 3 3 -73B) I35(ί)]-3-[(Azidiophenylacetyl)amino]-2-oxol-azetidinesulfonic acid, potassium salt A solution of 2.45 g of the above azetidinone and 3 g of monosilyltrifluoroacetamide in 20 ml of acetonitrile are kept for 1 hour at 40°C.

After cooling to 0°C the solution is treated with 1.88 g of trimethylsilyl chlorosulfonate and stirred for 5 hours under argon.

Finally 6.12 ml of a 2N solution of potassium-210 ethyl hexanoate in n-butanol are added and stirring is continued for 45 minutes. The solution is poured into 300 ml of ether and the precipitate is filtered off. A filtered solution of 1.2 g of precipitate in phosphate buffer IS (pH 5.5)is chromatographed on 100 ml of HP-20. Elution is performed with: 1) 20 ml buffer; 2) 200 ml HjO; 3) 200 ml wafer:acetone (9:1); 4) 200 ml wafer:acetone (3:1). The elution is monitored with TLC (Rydon test on SiO?). 25 ml fractions are taken and from fractions 15 and 16 280 mg of the title compound are obtained. A second column chromatography of this material yields 120 mg of the title compound, melting point 148°C, dec.

Method II: (S)-3-Amino-2-oxo-l-azetidinesulfonic acid, tetrabutylammonium salt (2.03 g; see Example 6A) and 0.9 g of (±)-o-azido-benzeneacetic acid are dissolved in 30 ml of acetonitrile and a solution of 1.03 g of dicyclohexylcarbodiimide in 10 ml 51333 of acetonitrile ie added. The temperature is maintained for one hour at 0°C and for 10 hours et rocn temperature. After filtering off the resultant precipitate, the solvent is distilled off In vacuo and the oily residue is dissolved in 20 ml of acetone and treated with 1.70 g potassium perfluorobutane sulfonate in acetone.

The addition of 10 al of ether crystallizes the title compound, melting point 149°C, dec.

Method III: a-Azidiophenylacetyl chloride (2.5 g) is added to a solution of 4.06 g of 3-*aino-2-oxo1-azetidinesulfonic acid, tetrabutylammonium salt (see Example 6A) end 5 g of propylene oxide in 30 ml of acetonitrile. After stirring for two hours the solvent is distilled off in vacuo and the oily residue is treated with one equivalent amount of potassium perfluorobutane sulfonate in acetone. After the addition of ether the title compound crystallizes and is filtered off, melting point 148-149°C dec.

Example 32 [3S (D) ] -3- ttlll (4-Methoxyphenyl)methoxy)carbonyl]amino] phenylacetyl] amino]-2-oxo-l-azetidinesulf onic acid, potassium salt To a solution of 2.03 g of 3-amino-2-oxo1-azetidinesulfonic acid, tetrabutylammonium salt (2.03 g; see Example 6A) and 1.58 g of D-a-I1((4methoxyphenyl Jmethoxy] carbonyl] amino] benzeneacetic -75acid in 50 ml of acetonitrile, a solution of 1.03 g of dicyclohexylcarbodiimide in 10 ml of acetonitrile is added dropwise. The temperature is maintained at 5°C for one hour and for 6 hours at room temperature. Separation.of the formed dicyclohexyl urea and distilling off the solvent yields the title compound as an oily residue. Treating this oil in acetone with potassium perfluorobutane sulfonate and ether yields 2.4 g of product, melting point 108-lll°C dec.

Examples 33-37 Following the procedure of Example 32, but substituting the compound listed in column I for (D)-o-I(I(4-methoxyphenyl)methoxy]carbonyl]amino]benzeneacetic acid, yields the compound listed in column II. 1 « 0 1 | | E 0 © © © © ι soI © © >» 1 >> 1 >. 1 © > 0 >« G 0 e c t . -* r* © M · e ι e » e 0 c * <ι « e *» . 0 1 1 «*> Sn MU 0 © «rt 4) · 0 © © x> •β —» -* -1 o| M ——Ό C C 00 X >iO © yl X X *0 fh ar ό « g X X 0 —»—* 1 0 X© acrt 4 φ ax© 4 *3·* •η x e o o w x © Q CM >« 0 x 0*0 >i 1 X M XI*» v r ο-* x a ** U fisf-rt X £ 0 X CM 0 or* ® E . *»r e — 40 1 ο a n e « ΟΙΝΕ· 54 so 1 *> X O D< — 0 X χ ~ 0 3 u X ri 4 9 u ϊ S ?7t> ·» t—F C — ο ON X 0 ~o X 0 I -HO • e ι Μφ 1 e* e~* H >, I -dH 0 e © m in V sh n e e-w-i a* 0 — >,0 *» >,XN ζ) B *rt 1 Μ Γ» E © 1 M w 1 E 1 «J SHJJUd β ο ι xx ι e o 0© soe© * a o * ι ** 0 0X0 e w 4 X X 1 0> 4 X X 1 —X 0« B C o 3 S ux C 0© **»» 9 0 cm **»©00© —f* o a. X X X X 9 0©© o "rtl Qtf. *-© ι a* — * I W ι r — · - μ 6 e e a *— >» — >i«M © "CM « r* ao s*» 1 *» 0 0 -Cl * -Cl * — οχτ> *» "SCdrt CM © ·—»*» 0» — Qrits X *— o f-»O x —ΛΟ—ι β — Xdfl X >© irt C — X 0-rt C — X 0© G ι μ e ο—* -gs·* a — C >3rt 1 U G O-rt ι ft e o © mt·*» Ο ι r x *» 1 0 X Μ X © 4 © 0 0 © 4© 4 o ios a rt *» F- χ go ©X 0 0© ioe a ι ο fi a —·—· X 0 1 Id X 90 ι a o e ο f-ι—· 4 U 3 o f— >|fw — E >«rt*f ·© >i 0 *rt e ri >ird© O' ri >mrt O' a xh c c — 1 C 1 art —» X frO U Xrt C G X X© c c — Q >0Ί +»< Of* XN +1 o Ort » * 8 >O-rt ** Q >1 O-rttor *>»* *» ——r 1 · 1 — X e X XωΧΧ*4ΧMX X *H XW *1 X —1 f* W— 1* O *» C* to 4J-rt «rt ** 4J ®rtrl . 4J 0 © © «η « o 9 · rt— X X Ort © 0 Ε M 0 © 0 o 9 0 <*) 0 0 9 0 — B · · — — o o ar* ~S«<« ~ Ε 4 M E — fi 4 M fi 1 rd 1 1 — 0 1 1 1 >1« r* f* ^irt > 0 X© 9*0 X 0 X-rt X© ox e n 0 0 0 0 0 X X 0 « X X X 4 X 0 X» o r-» X o 4J X 0 0 4 e « £S 0© Ε X 0 0 E-rt 0 4 E 0 ** c X 1 ** 0 *— X — c © 0 0 © 0 © 0 © 0 >x X 1 >1 4 >i U >iX c a X ri e e C 4 c a Φ 0 0 o 0 4 0 0 0 Q X© 6 C X κ X G X © ax 1 -rt ag a 0 ax Sx «* 6 >1*4 Sn >x X 1 ** 3 X 1 x e X 1 O CM «© ON 0 0 Q N X 1 — © X 1 XX X 1 4Jr-i — > X«-d x X ri 0 0 1 c 0 0 0 0 0 0 6 G 0 0 e c E G E G 1 © 1 X 1 © I *rt 1 © •e E o M*a -» fi E w E ** « e 0© — 4 — 0 **•0 - © ϋ o U^XN« — © E-* 4 *»© *— © *— © —' >1 3 > — >1 *- > ~ >1 1 C I X □ 1 C 1 c 1 G ο o CM 0 0 0 0 a o 1 J3 Ό 1 XX I X 1 X 1 X *0 — Ll *d rifi 0 «-* — M ri b© a χ o +10 0 +1 IQ X « X 0 o — 0 X 6 0 -r y — o - 0 0 0 © a E © NP m © P* « LJ © © © © © X U -77Example 38 (35)-3-1((((Methylthio)thioxomethyl]thio]phenylace tyl lami no] -2-oxo-l-azetldinesulfonie acid, tetrabutyl ananoniuro salt A solution of 1.03 g of (t)-a-(((methylthio)thioxomethyl]thio]benzeneacetic acid and 1.63 g of 3-amino-2-oxo-l-asetidinesulfonic acid, tetrabutylammonium salt (see Example 6A) in 30 ml of acetonitrile is treated with 0.8 g of dicyclohexyl10 earbodiimide dissolved in 10 ml of acetonitrile at -5®C. After stirring for about 16 hours the formed dieyclohexyl urea is filtered off and the mother liquor is evaporated. The remaining oily residue is chromatographed on a column of 400 g of silica (ethyl/acetate/methanol/water (8.5:1:0.5) is the eluent), yielding 1.3 g of the title compound Example 39 3(5)-3-(((( (Methylthio)thioxomethyl]thiojphenyl20 acetyl)amino]-2-oxo-l-azetidinesulfonic acid, potassium salt (35)-3-ί ί(((Methylthio)thioxomethyl]thioIphenylacetyl]amino-2-oxo-l-azetidinesulfonic acid, tetrabutylammonium salt (1.3 g; see Example 38) is dissolved in acetone and treated with an equivalent amount of potassium perfluorobutane sulfonate. The addition of ether crystallizes the title compound, which is filtered off, yield 0.18g of product, melting point 157°C, dec. 51333 -78Example 40 I3S(D)]-3-I[11 (4-Ethyl-2,3-dioxo-l-piperazinyl)carbonyl)amino]-2-thienylacetyl]amino]-2-oxo-lazetidinesulfonic acid, potassium salt (D)-e-11(4-Ethyl-2,3-dioxo-l-piperazinyl)carbonyl lamincg-2-thiopheneacetic acid (.3.25 g), 4.20 g of 3-amino-2-oxo-l-azetidinesulfonic acid, tetrabutylanenonium salt (see Example 6A) and 1.3 g of N-hydroxybenzotriazole are dissolved in 25 ml of acetonitrile. Dropwise addition of 2.06 g of dicyclohexylcarbodiimide dissolved in 10 ml of acetonitrile at -10°C takes 20 minutes and stirring is continued for about 16 hours, formed urea is filtered off, and the solvent is evaporated in vacuo. The remaining oil is dissolved in 50 ml of acetone and after treatment with an equivalent amount of potassium perfluorobutane sulfonate, the title compound crystallizes, melting point 185-187°C, dec.

Example 41 [3S(i)]-3-I(Bromophenylacetyl)amino)-2-oxo-lazetldinesultonic acid, potassium salt α-Bromophenylaeetyl chloride (1.4 g) in ml of acetonitrile is added dropwise to a solution of 5mM 3-amino-2-oxo-l-azetidinesulfonie acid, tetrabutylammonium salt, and 3 g of propylene oxide in acetonitrile at 0°C. After three hours stirring, the solvent is distilled off and the remaining oily residue is dissolved in 30 ml of acetone. The equivalent amount of potassium perfluorobutane sulfonate in acetone is added.

The addition of ether crystallizes the title -7910 Ο compound, melting point 135-137 C dec.

Example 42 [3S (i)]°3-[[I(Aminocarbonyljamino]-2-thienylacetyl]amino]-2-oxo-l-azetidinesulfonic acid, potassium salt Method 1; To a solution of 2 g of -(S)-3-amino-2-oxo-lazetidinesulfonic acid, tetrabutylammonium salt and 1.5 g of propylene oxide in acetonitrile is added 1.1 g of (±)-2-amino-4-(2-thienyl)-5(4H)oxazolone, hydrochloride. After three hours stirring af 0°C and one hour at room temperature, the solvent is distilled off and the oily residue is dissolved in 30 ml of acetone. By adding the equivalent amount of 'potassium perfluorobutane sulfonate in acetone, the title compound crystallizes out. Purification by column chromatography on HP-20 using water as eluent yields the title compound, melting point 218-222°C, dec. 80Method lit To a suspension of 2 g of (ί)-β-((aminocarbonyl ) amino] -2- thiopheneacetic acid, lOmM of (S)3-araino-2-oxo-l-azetidinesulfonic acid, tetrabutyl5 ammonium salt and lOmM of M-hydroxybenzotria2ole in 50 ml of acetonitrile at 0°C ia added (with stirring) lOmM of dicyclohexylcarbodiimide dissolved in 15 ml of acetonitrile. Stirring is continued for one hour at -15°C and for about 16 hours at room temperature. After filtering off the dicyclohexyl urea, the solvent is evaporated and the oily residue is dissolved in 50 *1 of acetone. Adding the equivalent amount of potassium perfluorobutane sulfonate in acetone yields crystalline product. Purification by column chromatography on HP-20, using water as an eluent, yields the title compound, melting point 220-223°C.

Example 43 (35(ί)]-3-([[[(Methylamino)carbonyl)amino]-2thlenylacetyljamino]-2-oxo-l-azetidinesulfonic acid, potassium salt (±) -a- [ [ (Methylamino) carbonyl] amino] -225 thiopheneacetic acid (0.54 g) and 1.00 g of (S)-3-amino-2-oxo-l-azetidinesulfonic acid, tetrabutylammonium salt (see Example 6A) are dissolved in 20 ml of acetonitrile and 0.5 g of dicyclohexylcarbodiimide is added at a temperature of 0°C. After stirring for 8 hours the dicyclohexylurea is filtered off and after distilling off -βΐthe solvent the oily residue is dissolved in 50 ml acetone and the equivalent amount of potassium perfluorobutane sulfonate is added. Crystalline product is filtered off and purification is performed on HP-20 using water as an eluent, melting point 205°C, dec.

Example 44 (3S (±)]—3— [[[ (aminooxoaeetyl) amino)-2-thienyl Xg acetyl]amino]-2-oxo-1-azetidinesulfonie acid, potassium salt Following the procedure of Example 42, method II, but substituting (±)-e-[{aminooxoacetyl)amino]-2-thiopheneacetic acid for (t)-c—[ (aminocarbonyl)amino]-2-thiopheneacetic acid, yields the title compound, melting point 218-222°C.

Example 45 [3S(R*)]-3-[ [ [ [ (4-Ethyl-2,3-dioxo-l-piperazinyl)20 carbonyl] amino] phenylacetyl ] amino]-2-oxo-l-azetidine sulfonic acid, potassium salt Following the procedure of Example 40, but substituting (R)-e-[[(4-ethyl-2,3-dioxo-l-piperazinyllcarbonyl]amino]benzeneacetic acid for (D)-α-[I(4-ethyl-2,3-dioxo-l-piperazinyllcarbonyl]amino]-2-thiopheneacetic acid, yields the title compound, melting point 155-157 C, dec. -82Example 46 3- (Acetylamino) -3-methoxy-2-oxo-l-azetidinesul fonic acid, potassium salt Method It A) 3- ((N-Acetyl-H-chloro) amino] -2-oxo-l-azetidinesulfonic acid, mixed eodium and potassium salt To a solution of 3- (acetylamino)-2-oxo-lazetidinesulfonic acid,potassium salt (172 mg, tee Example 23) in methanol (17 ml) containing 4% sodium borate decahydrate cooled to -15° to -10°C ia added tert-butyl hypochlorite (110 μΐ).

The mixture ia stirred in the cold for 1 hour, minutea, poured into 0.5 M monobasic potassium phosphate solution (50 ml), and the pB is lowered to 5.5. Solvent is removed in vacuo, and the residue is dissolved in a minimum amount of water and chromatographed on BP-20AG, 100-200 mesh (140 ml). Elution with water yields an oil (63 mg) which crystallizes on standing. Trituration with methanol/ether and then ether gives a solid (53 mg) melting point 124°C,slow dec. 51333 -03B) 3- (Acetylamino)-3-aethoxy-2-oxo-l-azetidinesulfonic acid, potassium salt 3-1 (N-Acetyl-N-ehlorofemino] "2-oxo-lazetidinesulfonic acid mixed salt (37 mg) is 5 dissolved in 1.5 ml of dry dimethylformamide and added to a solution of lithium methoxide (50 mg) in mettenol (2 ml) at -78°C. After stirring far 15 minutes at -78°C, a 0.5 M solution of monobasic potassium phosphate (10 ml) is added, and the solution is then acidified to pH 4 with IN hydrochloric acid.

To toe solution is added tetrabutylammonium hydrogen sulfate (70 ag) and the solution is extracted four times with methylene chloride. The combined extracts are dried (sodium sulfate) and solvent is removed in vacuo to give 55 mg of an oil.

Chromatography on 5.5 g of silica gel yields the oily product (41 mg, as the tetrabutylammonium salt (eluted with 8% methanol:92% methylene chloride). The oil (31 mg) is dissolved in water and passed through an ion-exchange column (5 ml of AG 50W-X2,K® form, 200-400 mesh, 0.6 meq/ml). Removal of wafer in vacuo gives an oil which crystallizes from methanol-ether. Trituration twice with ether gives the product as a colorless powder (11 mg); melting point 182-183°C, dec.

-BiMethod 11; λ) 3-Amino-3-»othoxy-2-oxo-l-azetidinesulfonic acid, tetrabutylammonium salt A 4* sodium borate decahydrate in methanol solution (100 pi) is added to a suspension of 10* palladium on charcoal (30 mg) in methanol (2 ml), and the mixture is stirred under an atmosphere of hydrogen for 15 minutes. 3-Methoxy2-oxo-3-[[(phenylmethoxy)carbonyl]amino]-ΙΙΟ azetidinesulfonic acid, tetrabutylammoniua salt (60mg; see Example 49) in methanol (2 ml) ie added and the mixture is vigorously stirred for 15 minutes under a hydrogen atmosphere. Catalyst is removed by filtration through Celite on a millipore filter (0.5 mu), and solvent is removed from the filtrate in vacuo, and the residue is extracted with methylene chloride. Removal of solvent under reduced pressure yields 35 mg of the title compound, as an oil.

B) 3-(Acetylamino)-3-methoxy-2-oxo-l-azetidinesulfonic acid, potassium salt To a solution of 3-emino-3-methoxy-2-oxo1-azetidinesulfonic acid, tetrabutylammonium salt (35 mg) in methylene chloride (10 ml) at 0°C is added propylene oxide (2 ml) and acetyl chloride (74 pi). After 2 hours the solvent is removed under reduced pressure and the residual oil is chromatographed on silica gel (4 g).

Elution with 6-8* methanol in methylene chloride gives an oil (18 mg) which is dissolved in water and passed through an ion-exchange resin (3 ml -85of AG 50W-X2, K form, 0.6 meq/ml). Removal of water from the eluate in vacuo yields the desired product (10 lag) .

S Example 47 N-(3-Methoxy-2°oxo-l-Bulfo-3-azetidinyl) -2-phenylacetamide, tetrabutylammonitna salt A) N-Chloro-N-(2-oxo-l-sulfo-3-azetidinyl)-3phenylacetanide, potassium salt To a solution of (S)-N-(2-oxo-l-sulfo-3azetidinyl)-2-phenylacetamide, potassium salt (50 mg; see Example 1) in methanol containing 4» eodium borate decahydrate (5 ml) cooled in a -5°C bath is added tert-butyl hypochlorite (20 pi). After stirring for 32 minutes, the mixture is poured into 0.5 M pH 5.5 potassium phosphate buffer at 0°C. The resultant solution (pH 5.9) is adjusted to pH 4.5, concentrated in vacuo to remove methanol, and chromatographed on HP-2QAG, 100-200 mesh (100 ml). After washing the column with 0.5 M pH 5.5 buffer (100 ml) and wafer the desired product is eluted with 9:1 water:acetone. Concentration in vacuo gives 50 mg of the title compound.

B) N-(3-Methoxy-2-oxo-l-sulfo-3-azetidinyl)-2phenylacetamide, tetrabutylammonium salt To a stirring solution of lithium methoxide (160 mg) in 5 ml of methanol cooled to -78°C is added a solution of N-chloro-N-(2-oxo-l-sulfo3-azetidinyl)~2-phenylacetamide, potassium salt 51333 -86(149 mg) in 10 ml of dry dimethylf ormamide. After addition is complete, the mixture is stirred for 15 minutes at -78°C, poured into 0.5 N monobasic potassium phosphate solution (100 ml), and washed three times with methylene chloride. Tetrabutylammoniw bisulfate (213 mg) is added to the aqueous layer, which is then extracted three times with methylene chloride. The combined extracts are dried (sodium sulfate), and solvent is removed in vacuo giving an oil (271 mg). Chromatography of the oil on silica gel (25 g) and elution with 41 methanol :96% methylene chloride yields 149 mg of the product as an oil.

Example 48 N-(3-Methoxy-2-oxo-l-sulfo-3-azetidinyl)-2phenylacatamide, potassium salt N- (3-Methoxy-2-oxo-l-sulf o-3-azetidinyl) 2-phenylacetamide, tetrabutylammonium salt (91 mg; see Example 47) is dissolved in water and passed through an ion-exchange column (10 ml of AS 50W-X2, X* form). The eluate is concentrated in vacuo and the residual oil solidifies on scratching with a methanol-acetone-ether mixture. After triturating twice with ether, the product is obtained as a solid (53 mg): υ 1762, 1665 cm1; NMR (CD,OD) 6 3.41 (S, 3H, OCH.), 3.59 (S, 2H, CH2), 3.84 (ABq, J-6.3 Hz, 2H, H4), 7.30 (m, 5H, aromatic).

Analysis: Cal. for C12H13^06.1/2H2O Calc.: C,39.88; H,3.62; 11,7.75 Pound : C, 39.62; H.3.65; N,7.60 51333 -87Example 4 9 3-Methoxy-2-oxo-3-I[(phenylmethoxy)carbonyl]amino)1-azetidinesulfonic acid, tetrabutylammonium salt Method I; S A) 2-0x0-3-(N-chloro-N-((phenylmethoxy)carbonyl]amino]-1-azetidinesulfonic acid, tetrabutylammonium salt (S)-2-Oxo-3-I[(phenylmethoxy)carbonyl]amino]-1-azetidinesulfonic acid, tetrabutylammonium salt (0.9g? see Example 4) dissolved in 80 ml of methylene chloride is added to a mixture (cooled to 0-5°C) of 3.17 g of sodium borate decahydrate and 11.8 ml of a 5.25% sodium hypochlorite solution in 70 ml of water. The reaction mixture is vigorously stirred for 55 minutes while cooling in an ice bath. After diluting the mixture with 0.5 H monobasic potassium phosphate solution, the product is extracted with methylene chloride (three 150 ml portions). Combination of the extracts, drying (sodium sulfate), and removal of solvent in vacuo yields the title compound as an oil (0.94 g).. 3-Methoxy-2-oxo-3- [ [ (phenylmethoxy) carbonyl] 25 amino]-1-azetidinesulfonic acid, tetrabutylammonium salt To a stirring solution of lithium methoxide (667 mg) in anhydrous methanol (10 ml) at -78°C is added a solution of 2-oxo-3-[N-chloro-N-[(phenyl' methoxy)carbonyl)arainoj-l-azetidinesulfonic acid, tetrabutylammonium salt (0.94 g) in dry dimethyl513 9 3 -88formamide (10 nl,. After stirring the mixture at -78°C for 1 hour, it is poured into 0.5 M monobasic potassium phosphate solution and extracted with methylene chloride (three 150 ml portions). The combined extracts are dried (aodium sulfate) and solvent is removed in vacuo to give en oil (0.83 g). The desired product is obtained by chromatographing the oil on silica gel (100 g, end eluting with 4-58 methanol- in methylene chloride to give en oil (513 mg): vaax(neat) 1767, 1720 cm"1» NMR (CDClj, δ 3.40 (S,OCH3), 3.03 (ABq, J-6.5 Hz, B4>, 5.08 (S,CB2), 6.00 (S,NH), 7.27 (S, aromatic).

Method lit To a solution of 3-benzyloxycarbonylamino3-methoxy-2-oxo-l-azetidinesulfonic acid, potassium salt (400 mg) in water ie added a 0.1 M tetrabutyl ammonium bisulfate solution (10.9 al, adjusted to pH 4.3 with potassium hydroxide). The mixture is extracted three times with methylene chloride, the extracts are combined, dried (Na^SOg) and solvent is removed in vacuo to give e foam (625 mg)r having spectral characteristics approximating those of the product of Method I. 51333 -89Example 50 3-Methoxy-2-oxo-3-1 ((phenylmethoxy)carbonyllamino] 1-azetidinesulfonic acid, potassium salt Method It A) 2-Oxo-3-[N-chloro-N-[(phenylmethoxy)carbonyl]amino]-1-azetidinesulfonic acid, potassium salt To a solution of (S)-2-oxo-3-II(phenylmethoxy) carbonyllamino]-1-azetidinesulfonic acid, potassium salt (1.00 g; see Example 3) in methanol (90 ml) containing 4* eodium borate decahydrate at -10°C is added tert-butyl hypochlorite (420 pi) After stirring for 2 hours at -10°C, 0.5 K monobasic potassium phosphate solution (100 ml) is added and the pH is adjusted to 6 with 1 N hydrochloric acid. After concentration of the solvent in vacuo to 30 ml, the aqueous solution is chromatographed on HP-20AG, 100-200 mesh (200 ml). After passage of a solution of monobasic potassium phosphate (50 g) in water (1000 ml), followed by water (2000 ml), the product is eluted with 10% acetone-90% water. Solvent is removed in vacuo and the title compound is crystallized from water to give a solid (530 mg), melting point 173-175°C. 51333 -908) 3-Methoxy-2-oxo-3-[[(phenylmethoxy)carbonyl]amino]-1-azetidinesulfonic acid, potassium «alt To a solution of lithium methoxide (874 rag) in dry methanol (10 ml) et -78°C is added 2-oxo5 3- [N-chloro-N- [ (phenylmethoxy) carb onyl J amino] 1-azetidinesulfonic acid, potassium salt (857 mg) in dry dimethylformamide (13 ml). After 15 minutes et -78°C the mixture is poured into 0.5 M monobasic potassium phosphate solution (200 ml) end the pB is adjusted to 5.5 with IN hydrochloric acid. The aqueous mixture is washed with methylene chloride (three 100 ml portions), and tetrabutylammonium bisulfate (1.159 g) is added. The product ia extracted vith methylene chloride IS (three 200 el portions), dried (sodium sulfate), filtered, and concentrated in vacuo. The residual oil is chromatographed on silica gel (150 g) and the product is eluted with 2-4% methanol in methylene chloride yielding the tetrabutylammonium salt of the product as an oil (701 mg). A portion of the oil (51 mg)is dissolved in water and passed through an ionexchange column (3 ml of AG 50W-X2, 200-400 mesh, K* form, 0.6 meq/ml). Concentration of the eluate in vacuo yields an oil (30 mg), which is crystallized by scratching with acetone: V (KBr, 1760, 1725 aa·1; NMR (D,O) 6 3.48 IQ&X * (S, 3H, OCHj), 3.92 (S, 2H, H4); 5.20 (S, 2H, CH2), 7.42 (S, 5H, aromatic)j m.p. 196-198·. -91Method II; A) l-Chloro-3- [N-chloro-N-[ (phenylmethoxy) carbonyl ] 2-azetidinone A solution of 3-1[(phenylmethoxy)carbonyl]5 amino)-2-azetidinone (440 mg.; see Example 20 in 40 ml of 4% methanolic borax is cooled to 0°C and 0.5 ml of t-butyl hypochlorite is added.

After 30 minutes at 0°C, the solution is poured into 200 ml of cold water and extracted with two ΧΘ 100 ml portions of ethyl acetate. The ^combined ethyl acetate layer is washed with water, dried, and evaporated in vacuo to give 546 mg of the title compound as an oil.

B) 3-Methoxy-3-[[(phenylmethoxy) carbonyl] amino]2- azetidinone A solution of 730 mg (0.0025 mole) of 1-chloro3- [N-chloro-N-[(phenylmethoxy)carbonyl]amino]-2azetidinone in 5 ml of tetrahydrofuran is cooled to -78°C and 4 ml of methanol containing 285 mg of lithium methoxide is added. After 20 minutes at -78°C, 0.6 ml of acetic acid and 0.6 ml of trimethylphosphite are added. The solution is stirred for 5 minutes at -78°C, allowed to warm to ambient temperature and stirred for 30 minutes.

The resulting solution is diluted with ethyl acetate, washed with 5% sodium bicarbonate,, water, 5% potassium bisulfate, water, saturated salt solution, and dried. Solvent removal gives an oil that is applied to four 20 x 20 cm xl mm 51333 -9210 silica gel plate*. Development with, benzeneethyl acetate (1:1) and isolation of the major UV-active band ol Rf-0.25 gives 91 mg of oil that crystallize* from ether to give a solid. Recrystallization from ether gives the title compound, melting point 112-114°C.

C) 3-Methoxy-2-oxo-3- [ I (phenylmethoxy) carbonyl] amino)-1-azetidinesulfonic acid, potassium salt λ solution of 25 mg of 3-methoxy-3-[[(phenylmethoxy) carbonyl]amino]-2-azetidinone in 0.175 ml each of dichloromethane and dimethylformamide is stirred for 24 hours with 55.4 mg of a complex of pyridine-sulfur trioxide. The resulting slurry is diluted with 5 ml of cold 0.5 M monobasic potassium phosphate (adjusted to pH 4.5) and extracted with ethyl acetate. The agueous layer is applied to a 40 ml HP-20 AG column. Elution with additional buffer, water, and water-acetone (9:1) gives 32 mg of the title compound as an oil that slowly solidifes. Crystallization from acetone gives tbe title compound, melting point 196-198°C, dec. -93Example 51 3-[[1,3-Dioxo-2-phenyl°3-(phenylmethoxy)propyllamino·] 3-methoxy-2-oxo-l-azetidinesulfonic acid, potassium salt A) 3-[(1,3-Dioxo-2-phenyl-3-(phenylmethoxy)propyl]amino)-3-methoxy-2-oxo-l-azetidine8ulfonic acid, tetrabutylammonium aalt Crude 3-amino-3-Biethoxy-2-oxo-l-azetidinesulfonic acid, tetrabutylammonium salt (431 mg, see Example 74), containing borax, is dissolved in 30 ml of dry acetonitrile. Dry pyridine ( 317 ul) is added and the solution stirred well at -10°C under dry nitrogen. o-(Benzyloxycarbonyl)phenylacatyl chloride (568 mg) in 3 ml of dry acetonitrile is added dropwise. Thin layer chromatography indicates the reaction to be complete after 15 minutes. Potassium phosphate buffer (0.5 M, pH 5.5, 17 ml) is added and most of the acetonitrile is removed in vacuo.‘ The residue is diluted with water and extracted three times with equal volumes of methylene chloride. The extract is dried over anhydrous sodium sulfate and evaporated in vacuo to give 1.032 g of crude product, which is dissolved in 3 ml of methylene chloride and chromatographed on a silica column using methylene chloridemethanol to give 470 mg of the title compound. -94B) 3-1 II,3-Dioxo-2-phenyl-3-(phenylmethoxy)propyl]amino]-3-methoxy-2-oxo-l-azetidinsulfonic acid, pota««ium aalt 3-[[1,3-Dioxo-2-phenyl-3-(phenylmethoxy)5 propyl] amino)-3-methoxy-2-oxo-l-azetidinesulfonic acid, tetrabutylammonium salt (470 mg) is dissolved in 15 ml of 30* acetone: water and put through a Dowex 50HX2 (K+foxm) column using the same solvent as eluent. The total eluate is evaporated in vacuo to yield 345 mg of an amorphous solid which is lyophilized to an amorphous powder, melting point 100-120°C.

Anal, for C^H^Ogl^SK Calc'd: C,49.37; 8,3.94 N,5.76; 6,6.59 Found: C,49.08; H,4.00; N,5.58; S,6.29 Example 52 (j)-3-11[(Cyanomethyl) thio]acetyl]amino]-3-methoxy2-oxo-l-azetidinesulfonic acid, potassium salt.3 To a solution of 3-amino-3-methoxy-2-oxol-azetidinesulfonic acid, tetrabutylansBonium salt (414 mg, see Example 74) in dry acetonitrile (50 ml) at -20°C ie added diethylaniline (210 pi) and cyanomethylthioacetyl chloride (169 mg).

After 10 minutes the solvent is removed under reduced pressure and a 0.1M solution of tetrabutylammonium sulfate (22.8 ml) adjusted to pH 4.3 with potassium hydroxide is added and the product extracted with three 50 ml portions of methylene 3® chloride, dried, filtered and concentrated under reduced pressure. The oily residue is purified on silica gel (60 g) and the product (294 mg) is eluted with 4* methanol-methylene chloride.

The purified product is passed through an ion -95exchange resin (16 ml AG 50W-X2(K+form), 0.6 mequiv./ml 200-400 mesh). Removal of water gives 164 mg of partially purified product.

The product is further purified on Diaion AG EF 20 (100 ml) using water as eluent. The solvent is removed under reduced pressure to give 126 mg of product which is triturated with ether to give 76 mg of the title compound, melting point 110-125°C.

Anal. Calc’d for CgEj^SjOgRs C,27.66; H,2.88; N, 12.10; 3,18.44 Found: C,27.25; H.3.00; N.10.84; S.17.53 Examples 53-56 Following the procedure of Example 42, Method II, but substituting the compound listed in Column I for (±)-a-f(aminocarbonyl)amino]-2thiopheneacetic acid, yields the compound listed in Column II.

Column I Column II -96- 1 Ch 1 Ch ι · e • c M C M 1 CM 1 v 0 M Xi Xi — c COH 0 Ό M o **» M m«h φ CM φ e μ ο o Ε A β M Xi £ m Ss m Gl 8 9 β · 1 Ό Ε χ» c —a M » In * 1 m«W V 4 9 0 W Η HX ** 4 a ** 0 □ c f* 0 IH fi X 1 M M 1 M M C «Μ M 4 Μ M A M 4 XM 4 XM 0 XM 9 X> • in c ι n Λ* β ζ) & Λ X»M u 0 00 e 9 A c 9 9 • X 8 A X S 0" e cp «Μ o c {»C! E ·+ ·+ m U Q 3 • 1 M 0 X«W Μ gjco » on a UN X Xi Μ XI 2 +>·+ +> 0 ι a 0 1 N 0 9 9m a ι +>+ c-j · C~ · ο ο η · NN · β Μ 0 X» c « « 2 ο ι η n e a g 8 β 0 ·, •rtrt C --, a a-+ fi. · • μ &υ| 8 XM * ·, . *" o · 8 ·· — a u ** 8 · a C Ό X» 0 "CHS U — a -o " 4 *Ol —- ·+ 1 ·+ S — —ιΌ et — trt A « — B 0 a «1 — ^-+O "MM » *3 β· — a χ n I NON ι xou "14 M » ι —, o a - n +i β ι «η xi 40 1 1 4 8JU nn 1+U I a r- f 9 m m ι nn go —, o oo ~ ϋ UN 1 1 ΜΜΙΛ -.+1 ι Sr* — a-+N F* 4 M fM F»m 1 n M *— β — Λ « <+ s * m e +1 0 0 π +10 0 -Η « Not! «no*; C X 4 1 — a β -ο ι — C «Η β — e a. e W-rt (0 0·+·+"+ M «+-+ ««HM m g ί om <*> χ e ο m nr a o nr s d " 4 fM 0.M — o a an -Q.no. — Q.n fi, 1 fM Ό M υ « I o 1 9 e « N c 1 OM 1 • XXI F-9 A 0 0 · 0 f s - *-ι ϋ c 0 M 0 4 M c s c « M fi 8 · § M § M a a M >9 ~ & X M X/ M 0 Xi X • XM • c o ΔΛ 0 Ό 0 • A XJ 4M A 0Ό • 1 0 0 U XM £ X 4 4' o u 0 1 0 O 0 4 Cr^ 0 0 0 c 4 0 CM c Μ o XC M xi Μ Ό Sm OM E 0 SM 4X1 ** E 4 0 4 O **· QJ " 4 —' 4 -* 4 " 5 1 — Φ I <0 " M ι e 1 0 o c ο χ ο QJ α m 1 4 1 xi 1 N 1 A u ** φ O C —a φ 4*1 9 4*1 0 Λ 0 « 0 *-* —* 4 ~A " 4e , . «η •f in to m m in m 51333 97Example 57 [3S (ί) )-3- [ [2°(Me thyl thio )-1-oxopropyl ] amino] -2oxo-l-azetidinesulfonic acid, potassium gait Following the procedure of Example 31, Method II, but substituting (i)-2-(methylthio)propanoic acid for (i)-a-azido-benzeneacetic acid, yields the title compound, melting point l73°C,dec.

Example 58 1Θ [3S (Ε·)ι) -3-(1((12,3-Dioxo-4-1 (phenylmethylene) amino] l-piperazinyl]carbonyl]amino]phenylacetyl]amino]-2oxo-l-azetidinesulfonic acid, potassium salt (R)-a-I[[2,3-Oioxo-4-((phenylmethylene)amino]1-piperazinyl]carbonyl]amino]benzeneacetic acid (0.7 g) and (S)-3-amino"2-oxo-l-azetidinesulfonic acid, tetrabutylammonium salt (0.8 g; see Example 6A) are dissolved in 20 ml of acetonitrile.

While stirring, 0.4 g of dicyclohexylcarbodiimide is added dropwise at 0°C. Stirring is continued for 18 hours and, after filtration, the solvent is distilled off leaving an oily residue. The residue is dissolved in acetone and treated with potassium perfluorobutane sulfonate to precipitate the title compound. Purification by column chromatography on HP-20 using water as eluent yields the title compound, melting point 193-194°C. -98Example 59 (35(2)1-3-(( (2-Amino-4-thiazolyl) [(2-me thoxy-2oxoethoxy)imino)acetyl]amino)-2-oxo-l-azetldineaulfonic aeid, potassium «alt (2)-2-Amino-o- ((2-methoxy-2-oxoethoxy)imino]4-thiazoleacetic acid (1.3 g) end (S)-3-amino-2-oxo1-azetidinesulfonic acid, tetrabutylammonium salt (2.03 g; aee Example 6A) is dissolved in 50 ml of acetonitrile and 1.03 9 of dicyclohexylcarbodiimide dissolved in 5 ml of acetonitrile is added dropwise at 0°C. After stirring for 15 hours and filtering off dieyclohexyl urea the solvent ie distilled off. The remaining oily residue is dissolved in acetone and treated with the equivalent amount of potassium perfluorobutane sulfonate. The title compound is isolated and purified by column chromatography on HP-20 using water as eluent, yielding the title compound melting point 195-198°C.

Example 60 (3S (R«) ]-3- [ [ [ [(3-[I(4-Chlorophenyl)methylene]amino] -2-oxo-l-imidazolidinyl ] carbonyl] amino] phenylacetyl ] amino] -1-azetidinesul fonic acid, potassium salt (S)-3-Amino-2-oxo-l-azetidinesulfonic acid, tetrabutylammonium salt (1.5 g; see Example 6A) in 100 ml of absolute diglyme (diethyleneglycoldimethyl ether), 1.5 g of (R)-a-(I(3-II(4chlorophenyl)methylene] amino]-2-oxo-l-imidazolidinyl ]carbonyl lamino] benzeneacetic acid, an equivalent amount of dicyclohexylcarbodiimide, and 0.5 g of 513 9 3 -99hydroxybenzotriazole are stirred together for 12 hours. The solvent ie removed in vacuo and the residue is dissolved in 50 ml of acetone and filtered. Acetone is distilled off and the residue is dissolved in 200 ml of methylene chloride. The solution is washed with aqueous sodium bicarbonate and then with aqueous sodium chloride solution. The methylene chloride layer is dried over sodium sulfate, evaporated to dryness and crystallized by the addition of ether. The compound is recrystallized from acetone-ether. The resulting white crystalline powder is dissolved in acetone and treated with aa equivalent amount of potassium perfluoro15 butane sulfonate in acetone. The title compound precipitates and is filtered off, yielding 1.4 g of product melting point 217-222°C.

Example 61 2Q [3S(R*)1-3-[11[12-0x0-3-[(phenylmethylene)aminol1-imidazolidinyl]carbonyl]amino]phenylacetyl]nmlno]-1-azetidinesulfonic acid, potassium salt (S) -3-Amino-2-oxo-l-azetidinesulfonic acid, tetrabutylammonium salt (2.25 g; see Example 6A) in 100 ml of dry dimethylformamide (DMF), an equivalent of dicyclohexylcarbodiimide, 2.5 g of (R)-α-I[[2-OXO-3-[(phenylmethylene)amino]-1imida zolidinyl]carbonyl] amino]benzeneacetic acid and 0.85 g of hydroxybenzotriazole are stirred together at ambient temperature for 12 hours.

After this time, DMF is removed in vacuo and the residue is dissolved in 50 ml of acetone. 51333 -100Th* precipitated urea ie filtered off and the mother liquor ie treated with an equivalent amount of potassium perfluorobutane sulfonate in 20 ml of acetone. After addition of 100 ml of ether, the title compound precipitates and is filtered off. Purification is achieved by dissolving the compound in DMF/acetone and precipitating with water, yielding 1.5 g of product, malting point 224-226°C, dec.

Example 62 [3S After the addition of ether, the title compound precipitates, yielding 2.0 g of material. The compound is purified by recrystallization from water and has a melting point of 240-245°C, dec. -101Sxample 63 [3S(Ra)]-3-[ (Hydroxyphenylacetyl)amino)-2-oxo-l azetidinesulfonic acid, potassium salt (S)"3-Amino-2-oxo-l-azetidinesulfonic acid, g tetrabutylammonium salt (1.5 g; see Example 6A) in 100 ml of dry dimethylformamide is stirred for about 16 hours with 1.5 g of dicyclohexylcarbodiimide, 0.5 g of hydroxybenzotriazole and 0.6 g of M-a-hydroxybenzeneacetic acid. 2.Q The solvent is removed in vacuo and the residue is dissolved in 20 ml of acetone. The precipitated urea is filtered off and the mother liquor is treated with an equivalent amount of potassium perfluorobutane sulfonate. After addition of 15 ether, the title compound precipitates and is filtered off, yielding 1.3 g of crude product.

The product is purified by chromatography using HP-20 and water/acetone (9:1) as eluent, and has a melting point of 145-150°C, dec.

Example 64 [3S(S*)]-3-I(Hydroxyphenylacetyl)amino]-2-oxol-azetidinesulfonic acid, potassium salt Following the procedure of Example 63, jg but substituting (S)-a-hydroxybenzeneacetic acid for (R)-o-hydroxybenzeneacetic acid, yields the title compound, melting point 195-197 C. -102Example 65 f35(ί)]-2-OXO-3-[(phenylsulfoacetyl) amino]-1azetidlnesulfonic acid, potassium «alt (1:2) (S)-3-Amino-2-oxo-l-azetidinesulfonic acid, tetrabutylaimnonium salt (2.25 g: sec Example 6A) in 100 ml of dry diglyme, 2.4 g of triethylamine end 0.3 g of dimethylaminopyridine are cooled to 0°C. (R)-a-(Chlorocarbonyl)benzenemethanesulfonic acid (1.8 g) in 20 ml of diglyme is added dropwise. The temperature is maintained for 2 hours, the solvent is distilled off in vacuo and the residue is dissolved in acetone. The insoluble precipitate is filtered off and the mother liquor is treated with an equivalent amount of potassium perfluorobutane sulfonate. After the addition of ether, the title compound precipitates and is filtered off, yielding 1.4 g of crude product; melting point 240-245°C dec., after recrystallization from water/acetone.

Example 66 [3S(Z)—3—[[(2-Amino-4-thiazolyl)[[(diethoxyphosphinyl)methoxy]imino]acetyl]amino]-2-oxol-azetidinesulfonic acid, potassium salt (S)-3-Amino-2-oxo-l-azetidinesulfonic acid, tetrabutylammonium salt (2.25 g; see Example 6A) in 100 ml of dry dimethylformamide is treated with 1.87 g of (Z)-2-amino-a-[[(diethoxyphosphinyl)methoxy]imino]-4-thiazoleacetic acid, 0.75 g of hydroxybenzotriazole and 2.29 g of dicyclohexylcarbodiimide for 12 hours with stirring. The precipitated urea is filtered off and the solvent -103removed in vacuo. The remaining oil is treated with an equivalent amount of potassium perfluorobutane sulfonate in 20 ml of acetone. After the addition of ether, the title compound precipitates and is filtered off, yielding 2.77 g of crude product. Purification of this crude product by column chromatography using HP-20 and water/ acetone (9:1) as eluent yields the title compound, melting point 155-160°C, dec.

Example 67 [3S(Z))-3-([(2~amino-4-thiazolyl)[(2-(1,1-dimethyle thoxy) -2-oxo-I-phenylethoxy] imino]acetyl]amino]-2oxo-l-azetidinesulfonic acid, potassium salt 15 (S)-3-Amino-2-oxo-l-azetidinesulfonic acid, tetrabutylammonium salt (2.25 g; see Example 6A) in 60 ml of dimethylformamide is stirred at roam temperature with 2.4 g of (2)-2-amino-a-I(2(1,1-dimethylethoxy)-2-oxo-l-phenylethoxy]imino]20 4-thiazoleacetic acid, 1 g of hydroxybenzotriazole and 1.5 g of dicyclohexylcarbodiimide for 12 hours. The solvent is removed in vacuo and the residue is dissolved in 50 ml of acetone. The precipitated urea is filtered off and the mother liquor is treated with an equivalent amount of potassium perflubrobutane sulfonate. After the addition of ether, the title compound crystallizes and is filtered off. Purification of the compound is achieved by HP-20 column chromatography using water/acetone (7:3) as eluent, yielding 1 g of product, melting point >250°C,dec. -104Example 68 (3S(Z)]-3-([(2-Amino-4-thiazolyl)((lH-tetrazol5-y lme thoxy) imino] acetyl] amino] -2-oxo-l-azetidinesulfonic acid, potaaaiua salt (S) -3-Amino-2-oxo-l-azetidinesulfonic acid, tetrabutylammonium eelt (1.9 g; see Example 6A) in 60 ml of dimethylformamide is treated with 1.4 g of (Z) -2-amino-a- [ (lH-tetrazol-5-ylmethoxy) imino]4-thiazoleaeetie acid, 0.7 g of hydroxybenzotriazole and 1.4 g of dicyclohexylcarbodiimide with stirring for 24 hours. After the solvent is removed in vacuo, the residue is dissolved in acetone and the precipitated urea filtered off. The mother liquor is treated with an equivalent amount of potassium perfluorobutane sulfonate in 10 ml of acetone.

The title compound is precipitated by the addition of 200 ml of ether. Purification is achieved by BP-20 column chromatography using HP-20 resin and water as eluent and yields 1.05 g of product, melting point 250°C, dec.

Example 69 [3S(Z)]-3-[I(2-Amino-4-thiazolyl) [(phenylmethoxy)imino] acetyl ] amino) -2-oxo-l-azetidinesul fonic acid, potassium salt (S) -3-Amino-2-oxo- 1-azetidinesulfonic acid, tetrabutylammonium salt (1.5 g; see Example 6A), 1.23 g of (Z)-2-amino-a-((phenylmethoxy)imino]-4thiazoleacetic acid, 0.57 g of hydroxybenzotriazole and 1.14 g of dicyclohexylcarbodiimide are stirred in 60 ml of dimethylformamide at room temperature for 24 hours. The precipitated urea is filtered off, the solvent removed and 51333 -105the residue treated with an equivalent amount of potassium perfluorobutane sulfonate in 10 ml of acetone. After the addition of 200 ml of ether, the title compound precipitates, is filtered S off and is purified hy HP-20 column chromatography using water/acetone (9:1) as eluent, yielding 1 g of material, melting point 200°C, dec.

Example 70 X0 [3S(Z)]-3-[I(2-Amino-4-thiazolyl)[(carboxymethoxylimino]acetyl]amino]-2-oxo-l-azetidinesulfonic acid, potassium salt (1:1) [3S(Z) ]-3-ίI(2-Amino-4-thiazolyl)[12-(diphenyl methoxy, -2-oxoethoxj5 imino] acetyl] amino] -2-oxo-l15 azetidinesulfonic acid, potassium salt (1:1) (1.3 g; see Example 30) is mixed with 5 ml of anisole.

At -15°C 25 ml of trifluoroacetic acid is added and the mixture is stirred for 10 minutes. Ether (100 ml) is added slowly at -10°C and subsequently 50 ml of petroleum ether. The precipitate is suspended with cooling in 20 ml of water and adjusted to pH 5.0 with diluted potassium hydroxide. The product is purified by chromatography on an HP-20 column, yielding 3.0 g of the title compound, melting point 230-235°C, dee. -106Exarople 71 [3S(2)]-3-Π(2-Amino-4-thiazolyl)J f2-oxo-2-(phenylmethoxy) ethoxy]imino]acetyl) amino)-2-oxo-l-azetidinesulfonic acid, potassium ealt Following the procedure of Example 28, but substituting (2)-2-amino-o-II2-oxo-2-(phenylmethoxy) ethoxy] imino]-4-thiazoleacetic acid for (Z)-2-amino-a-112-(diphenylmethoxy)-1,1-dimethyl2-oxoethoxy]imino]-4-thiazoleacetic acid, yields the title compound, melting point ca. 170°C, dec.

Example 72 [3S(2)]-3-[[[(2-Amino-2-oxoethoxy)imino](2-araino4 -thiazolyl) acetyl ] aminol-2-oxo-l-azetidinesulfonic acid, potassium salt Following the procedure described in Example 28, but substituting (2)-2-amino-o-[(2-amino2-oxoethoxy) imino]-4-thiazoleacetic acid for (Z)-2-amino-o-I[2-(diphenyImethoxy)-1,1-dimethyl2-oxoethoxy]imino]-4-thiazoleacetic acid, yields the title compound melting point 205-210°C, dec.

Example 73 (3S(2)]-3-[[(2-Amino-4-thiazolyl)(hydroxyimino)acetyl]amino]-2-oxo-l-azetidinesulfonic acid, potassium salt A solution of 0.6 grams of 90% hydroxybenzotriazole in 100 ml of dimethylformamide is stirred for one hour with 10 grams of 4A molecular sieves, filtered,and the filtrate added to a solution of 0.004 mole of (S)-3-amino2-oxo-l-azetidinesulfonic acid, tetrabutylammonium S1393 -107salt (see Example SA) in dimethylformamide. (2)-2-A»ino-a-(hydroxyimino)-4-thiazoleacetic acid (0.89 g) is added, followed by the addition of 0.91 g of dicyelohexylearbodiimide. The mixture is stirred for about IS hours, evaporated in vacuo and the residue dissolved in 20 ml of acetone and filtered. The addition of a solution of potassium perfluorobutane sulfonate causes the title compound to precipitate. Chromatography on HP-20 resin yields 0.44 g of product, melting point >240°C.

Example 74 3-Methoxy-2-oxo-3-[(2-thienylacetyl) amino]-1azetidinesulfonic acid, potassium salt A) 3-Amino-3-methoxy-2-oxo-l-azetidinesulfonic acid, tetrabutylammonium salt (+)_3-Methoxy-3-[[(phenylmethoxy)carbonyl]20 amino]-2-oxo-l-azetidinesulfonic acid, tetrabutylammonium salt (143 mg, see Example 49) is dissolved in 15 ml of dry methanol. Na^B^O^'lO HjO (12 mg, 0.1 equiv.) is added, followed by 10% palladium on carbon (72 mg). The mixture is hydrogenated at one atmosphere pressure for 15 minutes. The catalyst is removed by filtration and the filtrate evaporated in vacuo, yielding 114 mg of the title compound. -108B) 3-Methoxy-2-oxo-3-[ (2-thienylacetyl) aminoj-lazetidinesulfonic acid, tetrabutylammonium salt 3-Methoxy-3-araino-2-oxo-l-azetidinesulfonic acid, tetrabutylammonium salt (102 mg) is dissolved in 10 ml of dry acetonitrile. Dry pyridine (56.5 yl) is added and the solution is stirred well et -10°C under dry nitrogen. Thienylacetyl chloride (44 pi) in 1 al of dry acetonitrile is added dropwise. In 15 minutes the reaction is complete as shown by thin layer chromatography. Potassium phosphate buffer (0.5 M, pH 5.5, 4.2 ml) and tetrabutylammonium sulfate (8.5 mg, 0.1 eguiv.) ere added and most of the acetonitrile is removed in vacuo. The residue is diluted with water and extracted with three 20 ml portions of methylene chloride. The extract is dried over anhydrous sodium sulfate and evaporated in vacuo to yield 107 mg of a gum. The crude product is purified by chromatography on silica gel using methylene chloride-methanol, yielding 66 mg of the title compound.

C) 3-Methoxy-2-oxo-3- [ (2-thienylacetyl) amino] 1-azetidinesulfonic acid, potassium salt 3-Methoxy-2-oxo-3- [ (2-thienylacetyl) amino1-azetidinesulfonic acid tetrabutylammonium salt (154 mg) is dissolved in 3 ml of 30* acetone-water, passed through a column of Dowex 50W-X2 (K4- form) and eluted with the same solvent. The total eluate is evaporated in vacuo to yield 95 mg of product which is 51333 -109lyophilized to give an amorphous powder, melting point 120-13S°C.

Anal. Calc’d for C^Hj^OgS^: C.33.51; H.3.09; U.7.82; S.17.89 Found: C.33.46; H.3.08; N.7.92; 5,17.64 Example 75 (3S(Z)]-3-[[ (2-Amino-4-thiazolyl)[ (carboxymethoxy)imino] acetyl lamino] -2-oxo-l-azetidinesulfonic acid, potassium salt [3S(2))-3-(1(2-Amino-4-thiazolyl)ΐ[2-oxo-2(phenylmethoxy) ethoxy] imino ] acetyl] amino] -2-oxol-azetidinesulfonic acid, potassium salt (0.1 g; see Example 71) is dissolved in a mixture of ml of ethanol and 5 sal of water and hydrogenated at room temperature in the presence of 0.2 g of 10% palladium on charcoal. After 2 hours the catalyst is filtered off and the remaining solution is freeze-dried yielding the title compound. m.p. 235°(dec.).

Example 76 3- [ [ (S)- [ (Aminocarbonyl) asainoj-2-thienylacetyl]amino] -3-methoxy-2-oxo-l-azetidinesulfonic acid, potassium salt, isomer A To a solution of 3-amino-3-methoxy-2-oxo-lazetidinesulfonic acid, tetrabutylammonium salt (277 mg; see Example 46, method II, part A) in 15 ml of dry acetonitrile at -20°C is added pyridine (71 pi) 0.888 mmole) and (D)-2-amino-430 (2-thienyl)-5-(4H)-oxazoline hydrochloride (166 mg). The mixture is stirred for 10 minutes and a second -110aliquot of pyridine (71 ul) and oxazoline (166 mg) ia added. After another 10 minutes the solvent is removed under reduced perssure and the residue is dissolved in a water-acetone mixture and passed through an ion-exchange resin (20 ml AG 50W-X2,K-r fam, 200-400 mesh). Removal of the water from fractions 2- 3 gives a mixture of diastereomers (248 mg).

The product is purified and the diastereomers separated on Diaion AG HP 20 (130 ml) and eluted with water. Isomer A is eluted in fractions 23-28 (30 mg), and isomer B in fractions 35-45 (30 mg) (8 ml fractions). The middle fractions (10 mg) are combined with fractions from other runs and a total of 35 mg ot isomer A and 59 mg of isomer B is isolated.

Anal, for isomer A, m.p. 158-165*0.

Calc'd: ·Κ· 1/2H2O C, 31.05; H, 3.29; N, 13.18 FoundsC, 30.95; H, 2.97; II, 12.98 Anal, for isomer B, m.p. 160-170*C(dec).

Calc'd for CnH13N4O7S2-X«l/2H2O C, 31.05; H, 3.29; N, 13.18; S, 15.05 Found:C, 31.17; H, 3.09; N, 13.13; S, 15.09 Example 77 3- Methoxy-2-oxo-3-[(phenylsulfoacetyl)amino]-1azetidinesulfonic acid, dipotassium salt To a stirred solution of crude 3-amino-3methoxy-2-oxo-l-azetidinesulfonic acid, tetrabutylammonium salt (366 mg; see Example 74A) and 38 mg of borax in 35 ml of dry acetonitrile at -10eC under nitrogen is added 0.53 ml of dry pyridine followed by a two minute addition of a solution of α-sulfophenylacetyl chloride monoetherate -111(348 mg) in 8 ml ef acetonitrile. After 20 minutes the solvent is removed in vacuo and the residue is treated with 35 ml of 0.5 H pH 5.5 potassium phosphate buffer. Tetrabutylammonium hydrogen sulfate (383 mg) is added and the mixture is extracted three times with methylene chloride.

The methylene chloride is dried (sodium, sulfate) and evaporated to give 685 mg of crude product.

. The crude product is combined with 115 mg of crude from a second run and the combined material is purified on a column of SiliCAR CC-4 using methylene chloride and then 2,4,6,8 and 104 methanol in methylene chloride as eluent. The product, a mixture (about 6:1) of racemic diastereomers in the tetrabutylammonium salt form, is converted to the potassium salt by passage through Dowex 50W-X2 (K+ form) resin using 204 acetone in water as solvent. The product is lyophilized yielding 171 mg of the title compound, melting point 205-210°C, dec.

Anal. Calc'd for ci2H12N2°9S2K2'H2O: C,29.51; H,2.89 N,5.74; 5,13.10 Found: C.29.45; £,2.74; N.5.51; 5,12.82 Example 78 3-[(Carboxyphenylacetyl)amino]-3-methoxy-2-oxo1-azetidinesulfonic acid, dipotassium salt 3-[[1,3-Dioxo-2-phenyl-3-(phenylmethoxy)propyl) amino]-3-methoxy-2-oxo-l-azetidinesulfonic acid, potassium salt (39 mg; see Example 51) is dissolved in methanol (5 ml). Anhydrous potassium 51383 -112carbonate (3.9 mg) and 10% palladium on carbon (19 mg) are added and the mixture is hydrogenated at atmospheric pressure for 20 minutes. The catalyst is removed by filtration and the filtrate is evaporated in vacuo to yield * glassy residue (34 mg) which is lyophilized to an amorphous powder, melting point 178-190°C,dec.

Anal, for CuH12OgN2S Kj· 0.5 HjOs C,35.20; H,3.18: 11,6.32; 8,7.23 Found: C,35.51; H,2.96; N,6.29; S,6.92 Example 79 [3S (8)1-3-( [(2-Amino-4-thiazolyl)[[2-(1,1-dimethyle thoxy) -1-(methylthio)-2-oxoethoxy] imino] acetyl) — arainol-2-oxo-l-azetidinesulffonic acid, potassium salt Following the procedure of Example 73, but substituting (Z)-2-amino-a-[[2-(1,1-dimethy1ethoxy)-1-(methylthio,-2-oxoethoxy]imino]-4thiazoleacetic acid for (Z)-2-amino-a-(hydroxy20 imino)-4-thiazoleacetic acid, yields the title compound, melting point 130°C, dec. 51303 -113Example 80 (t)-3-Butoxy-3-(I(phenylmethoxy)carbonyl]amino]-2oxo-l-azetidlnesulfonic acid, potassium salt A solution of 185 mg of 2-oxo-3-[N-ehloro-N3 I(phenylmethoxy)carbonyl]amino]-l-azetidinesulfonic acid, tetrabutylammonium salt (see Example 49A) in 1 ml of dimethylformamide is cooled to -78°C and 0.73 N lithium n-butoxide (3.8 ml) in n-butanol is added at -78°C. After 15 minutes, 0.5 M monobasic potassium phosphate buffer is added and the product extracted info dichloromethane (three 40 ml portions), dried over sodium sulfate, filtered and concentrated in vacuo to give 179 mg of the corresponding tetrabutylammonium salt of IS the title compound.

To a solution of 109 mg of the tetrabutylammonium salt in acetone is added perfluorobutylsulfonic acid, potassium salt (60 mg) in acetone. The solvent is removed in vacuo and ethyl acetate is added. The product crystallizes and is collected and dried to yield 66 mg of the title compound, melting point 186.5-187.5°C, dec.

Example 81 [3±(E)]-3-Methoxy-3-([(methoxyimino) 12-(£(phenylmethoxy ) carbonyl] amino]-4-thiazolyl]acetyl]amino-2-oxo-l-azetidinesulfonic acid, potassium salt A suspension of 3-amino-3-methoxy-2-oxo30 1-azetidinesulfonic acid, tetrabutylammonium salt (Example 46, Method II, part A) (0.175 mmole) and sodium borate (0.0175 mmole) in 2 ml of -114diehlororaethane at 0°C is treated with 28 yl ot pyridine and 0.175 mmole of (E)-a-(methoxyinino)-2-[[(phenylmethoxy)carbonyl]amino]-4thiazolylacetyl chloride. After 1 hour, the mixture is diluted with dichloromethane and quenched with water. The organic layer is washed with water, saturated salt, dried, and evaporated in vacuo. The residue is purified on Mallinckrodt SilicAR CC-4 silica gel (20 g) to give 43 ng of the corresponding tetrabutylammonium salt of the title compound.

The tetrabutylammonium salt (43 ag) is dissolved in 0.5 ml of acetone and 20 ng of potassium perfluorobutane sulfonate in 0.5 ml of acetone is added. After addition of 3 ml of ether the solid is collected and dried in vacuo to give 28 mg of the title compound melting point 144-146°C, dec.

Example 82 I3t(Z)]-3-Methoxy-3-[I(methoxyimino)(2- [[(phenylmethoxy) carbonyl3 amino]-4-thiazolylJ acetyl]amino]2-oxo-l-azetidinesulfonic acid, potassium salt Following the procedure of Example 81, but substituting (Z)-a-(methoxyimino)-2-[I(phenylmethoxy) carbonyl]amino]-4-thiazolylaeetyl chloride for (E)-a-(methoxyimino)-2-(ί(phenylmethoxy)carbonyl]amino]-4-thiazolylacetyl chloride, yields the title compound, melting point 168-172°C, dec. -115Example 83 3-fl(R)-a-[1(4-Ethyl-2,3-dioxo-l-pipera2inyl)carbonyl lamino] phenylacetyl] am ine] -3-me thoxy-2oxo-l-azetidinesulfonic acid, potassium salt 5 To a stirred solution of 0.69 mmole of 3-amino-3-methoxy-2-oxo-l-azetidinesulf onic acid, tetrabutylammonium salt (Example 46, Method II, part A) in 30 ml of dry acetonitrile at -20°C under nitrogen is added 242 pi of dry pyridine followed by a solution of 352 mg of (R) - a-[ [ (4e thyl-2,3-dioxo-l-piperazinyl) carbonyl] amino] phenylacetyl chloride in 4 ml of acetonitrile.

After 1 hour 84 pi of pyridine is added followed by an additional 117 mg of the above named acid chloride in 1 ml of acetonitrile. The reaction is stirred for 20 minutes, diluted with 24 ml of 0.5 M pH 5.5 monobasic potassium phosphate buffer and concentrated in vacuo to remove acetonitrile. The agueous remainder is extracted three times with methylene chloride and the combined extracts are dried (Na^SO^) and evaporated leaving 546 mg of residue. Passage of the residue through a silica gel column using methylene chloride and then 2%, 4% and 6% methanol in methylene chloride, provides two fractions (285 mg and 173 mg) of the corresponding tetrabutylammonium salt of the title compound.

Passage of the 173 mg fraction through 4.5 g of Dowex 50-X2(K+) resin using acetone30 water as eluent yields 119 mg of the title compound. A 104 mg portion of this material is -116applied to a column of HP 20-AG resin in water. Sequential elution with water, 5* acetone in water and 104 acetone in water yields 60 mg of product as a mixture (ca. 1:1) of diastereomers. Lyophilization of the 60 mg fraction yields a solid, metling point 171-172°C, dec.

Example 84 N-(3-Butoxy-2-oxo-l-sulfo-3-azetidinyl)-2-phenylacetamide, tetrabutylammonium salt Following the procedure of Example 80, but subs tituting N-chloro-N-(2-oxo-l-sulfo-3-azetidinyl)2-phenylacetamide, tetrabutylammonium salt (see Example 47A for preparation of the corresponding potassium salt) for 2-oxo-3-[N-chloro-N-I(phenylmethoxy) carbonyl]amino]-1-azetidinesulfonic acid, tetrabutylammonium salt, yields the title compound as an oil: nmr (CDClg) 3.62 (s,2H,CgH5CH2), 4.03 (ABq,2H,v-7cps, C-4 CHj), 6.98 (s,lH,NH) and 7.30 ppm (S,5H,CgHs).

Example 85 (R)-3-Methoxy-2-oxo-3-t(phenylacetyl)amino]-1azetidinesulfonic acid, potassium salt A lM solution of dimethylformamide-sulfur trioxide complex is prepared by the slow addition of trimethylsilyl chlorosulfonate to dimethylformamide at 0°C followed by evacuation at 0.1 mm for 30 minutes at 0-25°C. Under an argon atmosphere, 50 mg of (R)-N-(3-methoxy-2-oxo-l-azetidinyl)phenylacetamide is dissolved in 0.2 ml of anhydrous dimethylformamide and cooled to 0°C. Cold 1M -117dimethylfonosside-Bulfur trioxide solution (0.428 ml) is added, the mixture is stirred for 2 hours and poured into 15 ml of 0.5 S monobasic potassium phosphate. The solution is extracted twice with dichloromethane (discard) aad 73 mg of tetrabutylammonium bisulfate is added. Extraction with dichloromethane (three 10 sal portions) gives a viscous oil after drying and evaporation in vacuo. Chromatography on Mallincrodt CC-4 silica gel (50:1)'using-2% methanol in dichloromethane .as eluant gives 34 mg of (R) -3-methoxy-2-oxo-3-l (phenylacetyl) amino] -1azetidinesulfonic acid, tetrabutylammonium salt. Ion exchange on Dowex 50W-X2 (K+, 10 equivalents) give’ the title potassium salt after lyophilization of the aqueous eluate: melting point 130°C, dec., [a]D=+52° (C=0.5, water,.

Example 86 (S)-3- [ [ [[(l-Ethyl-4-hydroxy-3-methyl-lH-pyrazolo[ 3,4-b] pyridin-5-yl)-carbonyl]amino]phenylacetyl]amino)-2-oxo-l-azetidinesulfonic acid, potassium salt Following the procedure of Example 73, but substituting a-(I(l-ethyl-4-hydroxy-3-methyl-lHpyrazolo [ 3,4-b] pyridin-5-yl) carbonyl] amino] benzeneacetic acid for (Z)-2-amino-a-(hydroxyimino)-4-thiazoleacetic acid, yields the title compound, melting point 233-236°C, dec. -118Example 87 (R)-3-Acetylamino-3-methoxy-2-oxo-l-azetidincaulfonic acid, potassium aalt A) 3-Acetylaraino-l- [l-carboxy-2-methyl(propyl)](3R)-3-«ethoxy-2-oxoazetidine To a solution of (6R-cis)-7-acetylamino-7methoxy-3-methyl-8-oxo-5-thia-l-azabicyclo [4.2.0]oct2-ene-2-carboxylic acid (650. mg) and sodiua bicarbonate (191 ag) in water is added a slurry (11 nl) of commercial grade Raney nickel (0.6 g/ml), whieh has been washed to neutrality with water.

The mixture is lowered into an oil bath preheated to 170°C and proceeds to reflux in 2-3 minutes, while maintaining the bath temperature at 150-170°C. After refluxing for 15 minutes, the reaction is quenched by cooling in an ice bath. Catalyst is removed by filtration through Celite.and the filtrate (pH 11) is adjusted to pH 2 with 1 N hydrochloric acid. After extracting the aqueous solution five times with ethyl acetate, the combined extracts are dried (Na^SO^) and solvent is removed in vacuo to give an oil (487 mg). Chromatography on silica gel yields the product (eluted in chloroform) as an oil (381 mg).

B) 3-Acetylaraino-l- [-l-(acetvloxy)-2-methyl(propyl)](3R)-3-methoxy-2-oxoazetidine The above azetidinone (464 mg) is dissolved in dry acetonitrile (15 ml) and the solution is purged with argon for 15 minutes. Copper acetate (359 mg) is added, stirred for one minute to dissolve the salt, and the lead tetraacetate 513 9 3 -119(797 mg) is added. While argon continues to bubble through the mixture, the temperature of the reaction is raised by lowering the flask into an oil bath, preheated and maintained at 55-65°C for 15 minutes. The mixture is allowed to cool to room temperature, filtered through Celite and the filter pad is washed well with acetonitrile. Solvent ie removed in vacuo from the combined filtrate and washings. The residue is taken up in water, and extracted four times with ethyl acetate. The combined extracts are dried (Na^SO^) and solvent is removed in vacuo to yield the desired product as an oil (382 mg).

C) (R)-N-(3-Methoxy-2-oxo-l-azetidinyl)acetamide The above oil is dissolved in a methanol (10 ml):wafer (1 ml) mixture, cooled in an ice-methanol bath at -10° to -15°C and potassium carbonate (194 mg) is added followed by sodium borohydride (53 ag). After Stirling at -15° to -8°C for 110 minutes, solvent is removed in vacuo, the residue is taken up in water and the solution is adjusted to pH 6 with 1 N hydrochloric acid.

Exhaustive extraction with ethyl acetate, drying (Na2SO4), and removal of solvent in vacuo yields an oil (224 mg). Chromatography of the oil on silica gel, eluting with 5% methanol:95% methylene chloride giving an oil (169 mg). The title compound crystallizes from ether-pentane to give 131 mg of material, melting point 106-112°C (sintering 103.5°C). -120D) (R)-3-Acetylamino-3-methoxy-2-oxo-l-azetidinesulfonic acid, potassium «alt Under an argon atmosphere, 50 ng of (R) -3-acetylamino-3-methoxy-2-oxo-l-azetidine 5 ie placed in a flask and cooled to 0°c. AIM solution of dimethylformamide-sulfur trioxide complex in dimethylformamide (.95 nl) is then added and the eolution is stirred for 15 ninutes.

The contents of the flask are then poured into 40 nl of 0.5 N XjHPO^ soultion and extracted twice with 10 al of nethylene chloride. Tetrabutylammonium sulfate (1.2 equivalents) is added to the aqueous solution and the resulting mixture is extracted with four 10 ml portions of methylene chloride. The extracts are then dried over NajSOj and concentrated to afford 39 mg of product. The tetrabutylammonium salt is converted to the title potassium salt by passing it through a column of Dowex 50-X2 (K+).

Concentration of the aqueous fraction gives mg of the potassium salt, with identical NMR spectra to that prepared in EX46Ib and by isolation from natural sources (Ex. 165). 513S3 -121Bxample 8§ (t)-3-[(Azidophenylacetyl)amino]-3-methoxy-2-oxD-lazetidinesulfonie acid, potassium gait A) (i)-3-ί(Azidophenylacetyl)amino]-3-methoxy-2oxo-l-azetidinesulfonic acid, tetrabutylammonium salt 3-Amino-3-methoxy-2-oxo-l-azetidinesulfonic acid tetrabutylammonium salt 1202 mgs; see Example 74A) is dissolved in 20 ml of dry acetonitrile.

To the well stirred solution at -20°C under dry nitrogen are added dry pyridine (167 pi) and a-azidophenylacetyl chloride (96 pi). After 20 minutes, 0.5M pH 5.5 monobasic potassium phosphate buffer (12 ml) is added and the acetonitrile removed in vacuo. The aqueous residue is extracted three times with methylene chloride.

The extract is dried over anhydrous sodium sulfate and evaporated in vacuo to give 281 mg of crude product as a gum. This is purified hy chromatography through a column of silica gel (30 g) using methylene chloride and mixtures of methylene chloride-methanol up to 6% methanol, and yielding 231 mg of the title compound.

B) (i)-3-[(Azidophenylacetyl)amino]-3-methoxy-»2oxo-l-azetidinesulfonic acid, potassium salt (i) — 3—[(Azidophenylacetyl)amino]-3-methoxy2-oxo-l-azetidinesulfonic acid, tetrabutylammonium salt (231 mg) in 30» acetone: water (15 ml) is passed through a column of Dowex 50W-X2 (K+ form; 3 ml) and eluted with water. The total eluate is evaporated in vacuo to give a colorless glass -122(168 mg) a· a 1:1 mixture of diastereomers.

Passage of this through a 60 ml column of HP20-AG using water and waterslOt acetone gives 71 mg of a 1:1 mixture of racemic diastereomers and 70 mg of a 1:3 mixture of racemic diastereaners. The 1:1 mixture is lyophilized and dried in vacuo at 40°C to give the dried product as a hemihydrate, melting point dec. 130°C.

Analysis for εχ2^.2Ν5°6*Κ’ *5 H20: Ca^c'^: c' 35.90; B, 3.26; N, 17.45; S, 7.98 Found: C, 35.94; B, 3.07; N, 17.24; S, 8.02 Example 89 3-1 (Azidophenylacetyl)amino]-3-methoxy-2-oxo-lazetidinesulfonie, potassium salt, isomer A The 1:3 mixture of racemic diastereomers obtained in Example 88B is allowed to stand in deuterated water at room temperature, and isomer A crystallizes. After refrigeration, the mother liquor is removed, and the crystals (28 mg) are dried in vacuo at 40°C, melting point 130°C, dec, as a ποηο-deuterate.

Example 90 13± (R*) 3 -3* [ΙΓΙ (4-Ethyl-2,3-dioxo-l -piperazinyl) carbonyl ] amino 1 phenylacetyl] amino] -3-methoxy-2oxo-l-azetldinesulfonlc acid, potassium salt To a stirred solution of 3-amino-3-methoxy2-oxo-1-azetidinesulfonic acid, tetrabutylammonium salt (0.69 mmol; see Example 74A) in 30 ml of dry acetonitrile at -20°C under nitrogen is added 242 yl of dry pyridine followed by a solution of 51333 -123352 mg of (R)-ο-ίI(4-ethyl-2,3-dioxo-l-piperazinyl) carbonyl]amino]phenylacetyl chloride in 4 ml of acetonitrile. After 1 hour, §4 μΐ of pyridine ia added followed by 117 mg more of the acid § chloride in 1 ml of acetonitrile. The reaction is stirred for 20 minutes, diluted with 24 ml of 0.5 M pH 5.5 monobasic potassium phosphate buffer, and concentrated in vacuo to remove acetonitrile. The agueous remainder is extracted three times with methylene chloride and the combined methylene chloride extract is dried (NajSO^) and evaporated to a residue (546 mgl. Passage of this material through a column of SilicAR CC-4, using methylene chloride and then X5 2%, 4%, and finally 6% methanol in methylene chloride provides two fractions (2S5 mg and 173 mg) of purified product in the tetrabutylammonium salt form.

Passage of the 173 mg portion through 4.5 g 2Q of Dowex 50-X2 (K ) resin using acetone-water yields 119 mg of potassium salt.' A 104 mg portion of this material is applied to a column of HP20-AG resin in water. Sequential elution with water, % acetone ia water, and finally 10% acetone 2g in water provides 60 mg of product as a mixture (ca. 1:1) of diastereomers and 21 mg of product as a mixture (ca. 9si) of diastereomers. Lyophilization of the 60 mg fraction yields the title compound, melting point 171-172°C, dec. 3Q Analysis for ClgH22N5OgSK-H2O: Calc’d: C, 41.23; H, 4.37; N, 12.65; S, 5.78 Found: C, 41.39; H, 4.12; N, 12.58; S, 5.63 51333 -124Lyophilization of the 21 mg fraction yield· the title ccopound, melting point 171-172°C, dec. Analysis for C^jHjjNgOjSK’HjO: Calc'd: C, 41.23 B, 4.37; N, 12.65 Found: C, 41.43; B, 4.11; N, 12.28 Treatment of the 285 mg fraction of tetrabutylammonium salt with Dowex 50-X2(K+) provides 145 mg of potassium salt, which is combined with, the remaining 15 mg of the aforementioned 119 mg portion of Dowex resin derived potassium salt. Passage of this materiel through HP2Q-AG, as already described, provides an additional 31 mg of product as a mixture (ca. 1:1} of diastereomers and an additional 42 mg of product as a mixture (ca. 9:1) of diastereomers. The total amount of (1:1) mixture of diastereomers is 91 mg, and the total amount of (9:1) mixture of diastereomers is 63 mg.

Example 91 [3S(Z)]-3-I((Methoxyimino)[2-(I(phenylmethoxy)carbonyl]amino)-4-thiazolyl]acetyl] amino]-2-oxol-azetidinesulfonic acid, potassium salt To a solution of (S)-3-amino-2-oxo-lazetidinesulfonic acid, tetrabutylammonium salt (0.170 mmol; see Example 6A) and sodium borate (0.170 mmol) in 2 ml of methylene chloride at 0°C is added pyridine (62 ul) and (Z)-e-(methoxyimino) -2-[[(phenylmethoxy)carbonyl] amino]-4thi&2oleacetyl chloride (0.51 mmol). The reaction mixture, after 40 minutes, is diluted with methylene chloride and water, followed by 0.1 M 51333 -125tetrabutylammonium sulfate buffered to pH 4 (5.1 ml). The organic layer is separated and washed with water, adjusted to pH 2, water adjusted to pH 7, water saturated with sodium chloride, dried over sodium sulfate, filtered, and concentrated in vacuo. The residue is purified on SilicAH CC-4 silica gel 110 g) and the product eluted with 10% methanol/aethylene chloride.

The tetrabutylammonium salt, after dissolution in acetone/water, is passed through an ion-exchange resin (8 ml, AG 50W-X2, K+ form 100-200 mesh.). Removal of the water in vacuo from fractions 1-2 give 40 mg of the title compound, melting point 172-174°C, dec.

Analysis for C^jH^gN^OgSjK-HjO: Calc'd: C, 37.84; H, 3.33; N, 12.99; S, 11.87 Found: C, 37.95; H, 3.30; N, 12.73,- S, 11.53 Example 92 (i)-3-Butoxy-2-oxo-3-[(phenylacetyl)amino]-1azetidinesulfonic acid, potassium salt A) 3- [Chloro(phenylacetyl) amino]-2-oxo-l-azetidine25 sulfonic acid tetrabutylananonium salt A solution of (S)-2-oxo-3-[ [phenylacetylamino]-1-azetidinesulfonic acid, tetrabutylammonium salt (350 mg) in methylene chloride (3 ml) is added to a suspension 3Q of sodium borate (1.27 g) in a 5.25% solution of sodium hypochlorite (4.72 ml) and water (20 ml) -126at 0°C. After 1 hour 0.5 H monobasic potassium phosphate (25 ml) is added and the mixture is extracted three times with methylene chloride (50 ml portions). The organic extracts are dried over sodium sulfate, filtered and concentrated in vacuo to give 344 mg of the title compound. Β) (+)-3-Butoxy-2-oxo-3-I (phenylacetyl) amino]-1azetidlnesulfonic acid, tetrabutylammonium salt A solution of 3-[chlorophenylacetylamino]-1-azetidinesulfonic acid, tetrabutylammonium salt (344 mg) in dimethylformamide (5 ml) ie added to 0.73 N n-lithium butoxide in n-butanol (6 ml) in dimethylformamide IS (1 ml) at -78°c under an inert atmosphere. After 10 minutes the mixture is diluted with 0.5 M monobasic potassium phosphate solution'(175 ml). After extracting three times with methylene chloride, the organic extract is dried over sodium sulfate, filtered, and the solvent' removed in vacuo. The residue is purified on SilicAB CC-4 silica gel (80 g) and the title canpound (130 mg) is elutpd with 4-8% methanol in methylene chloride.

C) (i)-3-Butoxy-2-oxo-3-[(phenylacetyl) amino]1-azetidinesulfonic acid,' potassium salt (±)-3-Butoxy-2-oxo-3-[(phenylacetyl) amino)1-azetidinesulfonic acid, tetrabutylammonium salt (43 mg) is dissolved in a water-acetone mixture (9:1) and placed on a cation exchange column (Dowex AGMP 50W-X2, 100-200 mesh, 5 g, K+ foxm). 513 93 -127The product is eluted with water and the eulate concentrated in vacuo to give 20 mg of the title compound, melting point 122-125°C.

Analysis calc’d for CjgHjgNjOgSK-1/2 H2O: C, 44.66; H, 4.96; N, 6.95; S, 7.94 Found: C, 44.77; H, 4.76; N, 6.76; S, 7.75 Example 93 (i) -3-Ethoxy-2-oxo°3-I (phenylacetyl) aroinol-110 azetidinesulfonic acid, potassium salt 3-lChloro (phenylacetyl) amino] -2-oxo-lazetidinesulfonic acid, tetrabutylammonium salt (200 mg see example 92A) in dimethylformamide (4 ml) is added to 0.5 N lithiixD ethoxide in ethanol (12.20 ml) at -78°C under an inert atmosphere. After 10 minutes the mixture is diluted with 0.5 K monobasic potassium phosphate solution (15 ml).

After extracting three times with methylene chloride, the organic layer is dried over sodium sulfate, filtered and the solvent removed in vacuo. The residue is purified on SilicAR CC-4 silica gel (20 g) and the tetrabutylammonium salt of the product (40 mg) is eluted with 21 methanol in methylene chloride.

The tetrabutylammonium salt is dissolved in a water-acetone mixture (9:1) and placed on a cation exchange column (Dowex AGMP 50W-X2, 100-200 mesh (5 g, K form)). The product is eluted with water and the eluate concentrated in vacuo to give 25 mg of the title compound, melting point 94-96°C.

Analysis calc'd for C^jH^NjOgSK: C, 42.62; H, 4.10; H, 7.65; S, 8.74 Found: C, 40.36; H, 3.66; N, 6.77; S, 8.44 -128Example 94 I3i(Z)]-3-(] (2-Amino-4-this zolyl) (me thoxy imino) •cetyl]amino]-3-methoxy-2-oxo-l-azetldlnesulfonic acid, potassium aalt 3-Amino-3-methoxy-2-oxo-l-azetidinesulfonic acid, tetrabutylammonium salt (see Example 46, method XX, part λ) ia dissolved in acetonitrile (20 al) and pyridine (1 al) and added to a vigorously stirring suspension of (Z)-a-(methoxy10 imino) -2-aaino-4-thiazolbacety.l chloride in acetonitrile (20 al) cooled to d-5°C. After stirring cold for 1 hour the mixture is diluted with 0.5 M monobasic potassium phosphate solution (100 al) (pH of the mixture ie 4.8) and solvent is removed in vacuo. The residue is taken up in a minimal amount of water containing a small amount of acetone. Chromatography on ion exchange resin (AG 50W-X2, 100-200 mesh, K+ form 200 ml) gives the crude product as the potassium salt upon elution with water. Further purification on HP-20 resin (200 al) using water as the eluant gives 59 mg of the product as a powder after trituration with acetonitrile-ether and then twice with ether. The product is an amorphous powder that aelts slowly and decomposes above 150°C.

Analysis calc'd for C1(jH12N5O7SK: C, 28.77; H, 2.90; N, 16.78; S, 15.36; K, 9.37 Found: C, 27.77; H, 2.82; N, 15.87; S, 13.63; K, IQ.11 -129gxample gs (3R(R*) and 3S(Sa)]-3-III(aminoearfconyllaaBina]phenylacatyl)amino]-3-methoxy°2-oxo-l-azetidineeulfonie acid, potaaaium salt 5 Α) (ί)-3-[ (Aminophenylacetyl) amino]-3-methoxy-2oxo-l-azetidinegulfonie acid, inner gait (+) -3- {(Azidophenylacetyl) amino] -3-aethoxy2-oxe-l-azetidinesulfonic acid, potassium salt 10 (209 mg; see Example 88)iis dissolved in 6G sal of dry methanol. Anhydrous trifluoroacetic acid (0.6 ml) and 10Q palladium on carbon (105 mg] arc added and the mixture is hydrogenated for 1 hour. The catalyst is removed by filtration and the filtrate is evaporated in vacuo to yield 271 mg of crude product. 8) ]3R(R*) and SStS^J-S-KKAminocarbonyDamino]phenylacetyl] amino]-3-methoxy-2-oxo-l-azetidine20 sulfonic acid, potassium salt (+)-3-1 (Aminophenylacetyl) amino]-3-methoxy2-oxo-l-asetidinesulfonic acid, potassium salt (271-mg) is dissolved in 6.5 ml of water.

Potassium cyanate (87 mg) is added snd the mixture is stirred at room temperature for 3 hours.

The solution is concentrated in vacuo to approximately 2 ml and chromatographed on a 100 ml column of HP20-AG using water as eluant. Isomer A (29 mg) is isolated after lyophilization, melting point 160°C, dec.

Analysis for C^gH^jN^O^K monohydrate: Calc'd: C, 36.44 H, 3.99; N, 13.07; S, 7.48 Found: C, 36.35; H, 3.79; N, 12.81; S, 7.32 51333 -130Example 96 [3R(S*) and 3S(R*)]-3-in(Aminocarbonyl)amino)phenylacetyl]amino]-3-methoxy-2-oxo-l-azetidinesulfonic acid, potassium salt 5 Along with I3R(R*) and 3S(S*)J-3-[II(aminocarbonyl) amino]phenylacetyl]amino-3-methoxy-2ozo-1-azetidinesulfonic aeid, potassium salt produced in Example 95, there are produced 21 mg of I3R(S*1 and 3S(8*11-3-1II(aminocarbonyl110 amino] phenylacetyl] amino]-3-me thoxy-2-oxo-lazetidinesulfonic acid, potassium salt as a lyophilate, melting point 16Q°C dec.

Analysis for C^gH^jK^OySK sesquihydrate: Calc'd: C, 35.69; H, 4.14; M, 12.81; S, 7.33 Found: C, 35.98; 8, 3.87; N, 12.50; S, 7.32 Example 97 [3—(S*)]-3-Methoxy-3-IIII[2-oxo-3-I(phenylmethylene)amino]-l-imidazolidinyl]carbonyl]amino]-2-thienyl20 acetyl]amino]-2-oxo-l-azetidinesulfonic acid, potassium salt To a stirred solution of 3-amino-3-methoxy2-oxo-azetidinesulfonic acid, tetrabutylammonium salt (306 mg of crude material assumed to contain 25 274 mg of organic material, prepared as described in Example 74) in 20 ml of dry acetonitrile at -20°C under nitrogen is added 0.30 ml of dry pyridine (3.72 mmol) followed by 484 mg of (S)-I[[2-oxo-3-pheny lmethylene J-amino ]-1-imidazolidinyl]carbonyl]amino)-2-thienylacetyl chloride partially dissolved and suspended in 10 ml of dry acetonitrile. 513 0 3 -13110 The reaction is stirred and allowed to rise to 0°C over the course of 1 hour.

The reaction was diluted with a large volume of methylene chloride and then treated with 22 ml of 0.5 M pH 5.5 monobasic potassium phosphate buffer. The aqueous layer is washed with methylene chloride and the combined methylene chloride extract is washed with water, dried (NajSO^), and evaporated to a residue (508 mg). This residue is chromatographed on 5Q g of SilicAR CC-4, using methylene chloride and then 28 and 48 methanol in methylene chloride to give 251 mg of the tetrabutylammonium salt of toe title compound.

To this salt (251 mg) in acetone is added a solution of 107 mg of perfluorobutanesulfonic acid potassium salt in several milliliters of acetone. Ethyl acetate is added, and the precipitate is washed three times with ethyl acetate by centrifugation, and dried in vacuo at 40°C/lmm for 2 hours to give 95 mg of desired potassium salt as a mixture (ca. 1:2) of diastereomers having a melting point 200°C, dec.

Calc'd for C2lH21NS°8S2Ks C42’85'’ H' 3.60; X, 14.28; S, 10.87 Found: C, 43.02; H, 3.74; N, 13.94; S, 10.71 51333 -132Example 98 (-—cis) -4-Hethyl-2-oxo-3-I [ (phenylmethoxy) carbonylj amino]-1-azetidinerulfonic acid, potassium salt A) N-Benzyloxy-t-boe*'-allothreonine amide A solution of 6.9 g of d,l-t-boc-allothreonine and the free amine from 5.3 g of o-benzylhydroxy1amine*BCl (*v0.033 mole, ethyl acetate-sodium bicarbonate liberation) in 80 ml of tetrahydrofuran is treated with 4.82 g of N-hydroxybenzotriazole and 6.5 g of dicyclohexylcarbodiimide in 20 ml of tetrahydrofuran. After stirring for about 16 hours at room temperature the slurry is filtered, concentrated in vacuo and chromatographed on a 400 ml of silica gel column. Elution with -10% ethyl acetate in chloroform gives 6.8 g of the title compound in fractions (200 ml each) 7-22.

B) ( ±— ci^-N-Benzyloxy-3-t-butoxycarbonylamino-4methylazetidinone A solution of 6.8 g of N-benzyloxy-tboc-allothreonine amide in 200 ml of tetrahydrofuran is stirred for about 16 hours with 5.24 g of triphenylphosphine and 3.2 ml of diethylazodicarboxylate. The solvents are evaporated in vacuo and the residue is chromatographed on a 500 ml silica gel column. Elution with methylene chloride followed by crystallization from ether gives a total of 2.65 g of azetidinone. Bechromatography of the mother liquors and mixed fractions give an * boc is used to describe butoxycarbonyl -133additional 0.6 g. Crystallization of a portion o twice from ether (-20 C) gives the analytical sample of the title compound melting point 14O-142°C.

Cl (j~cis)-3-t-Butoxycarbonylamino-l-hydroxy-4methylazetidinone A solution of 3.2 g of cis-N-benzjtloxy 3-t-butoxycarbonylamino-4-methylazetidinone in 200 ml of 95% ethanol is stirred in an atmosphere of hydrogen with 0.7 g of 10% palladium on charcoal. After 40 minutes the slurry is filtered (uptake 249 ml) and the filtrate is evaporated and triturated with ether to give, in two crops, 2.05 g of solid, melting point 134-136°C.

D) (+—cis-3-t-Butoxycarbonylamino-4-methyla2etidinone A solution of 2.05 g of cis-3-t-butyloxycarbonylamino-1-hydroxy-4-methylazetidinone in 60 ml of methanol is treated with a total of 9o mi Of 4.5 m ammonium acetate (40, 20 and 30 ml portions) and 45 ml of 1.5 M titanium trichloride (20, 10 and 15 ml portions] the second and third additions are made after 15 and 120 minutes, respectively. After 135 minutes the solution is diluted with an equal volume of 8% sodium chloride and extracted with three 300 ml portions of ethyl acetate. The combined organic layer is washed with a mixture of 100 ml each of 5% sodium bicarbonate and saturated salt, dried, and evaporated. Trituration with ether gives, in two crops, 1.65 g of solid. A portion of the first crop is recrystallized from ether to give the analytical sample, melting point 176-178.5°C. 51333 -134E) ( i -cis)-3-Benzyloxycarbonylamino-4-ineth.ylazetidinanc A eolution of 1.55 g of cis-3-t-butoxycarbonylamino-4-methylazetidinone in 4 al each of methylene chloride and anisole is cooled to 0°C and 50 ml of cold trifluoroacetic acid is added.

After 90 minutes the solvents are evaporated in vacuo (benzene added and evaporated three times].

Tbe residue is dissolved in 25 ml of acetone, the initial pH (2.51 is raised to 7 with 58 sodium bicarbonate, and 2 ml of benzylchlorofornate is added. The solution is kept at 0°C and pH 7 for 4 hours and the acetone is removed in vacuo to give a slurry that is filtered. Tbe filtrate is saturated with salt and extracted with methylene chloride. The solid is dissolved in methylene chloride and dried. The organic layers are combined, concentrated, and the residue chromatographed on a 200 ml silica gel column. Elution with 3:1 chloroform, ethyl acetate gives 850 mg of the title compound in fractions (100 ml each) 4-11. Crystallization of a small sample from ether gives the analytical sample, melting point 165-166°C.

F) (j-cis) -4-Methyl-2-oxo-3- [ I (phenylmethoxy) carbonyl]amino]-1-azetidinesulfonic acid, potassium salt To a suspension of Cis-3-benzyloxycarbonylamino-4-methylazetidinone (0.75 g) in 7 ml each of dimethylformamide (dried with 4A sieves activated at 320°C for 15 hours under argon flow) and methylene chloride (dried through basic ΑΙ^,Ο^) 51333 -135is added 1.66 g of pyridine-eulfur trioxide complex. After 3 hours otirring at room temperature under nitrogen, an additional amount of pyridine-sulfur trioxide complex (1.66 g) is added. The reaction S mixture is then stirred at room temperature under nitrogen for about 16 hours. The dimethylformamide is removed in vacuo to give 4.6 g of residue which is dissolved in 300 ml of 0.5 M monobasic potassium phosphate solution C40°C for -15 minutes). The solution is cooled, passed through a column of HP-20 resin (3 cm x 60 cm) with 400 al of 0,5 H monobasic potassium phosphate 1Lof distilled water and C14:l) water:acetone to give 280 mg of product in fractions 13 to 26 (1QQ ml each). Crystallization from MeOH: petroleum ether gives 757.5 mg of an analytical sample, melting point 214-215.5°C, dec.

Analysis calc'd for C^jH^NjSOgK: C, 40.90; H, 3.72 N, 7.95; S, 9.10; K, 11.10 Found: C, 40.43; H, 3.60; N, 7.89; S, 8.69; K, 10.82 Example 99 (3S-trans) -4 -Methyl-2-oxo-3 11 (phenylmethoxy) carbonyl] 25 amino)-1-azetidinesulfonic acid, potassium salt Following the procedure of Example 98, but substituting 1-t-boc-threonine for d,l-t-bocallothreonine, yields the title compound, melting point 133-135°C.

Analysis calc'd for C^jH^NjOgSK: C, 40.90; H, 3.72; N, 7.95; S, 9.10; K, 11.10 Found: C, 40.72; H, 3.60; N, 7.99; S, 8.80; K, 10.82 -136Ixample 100 C3S-tram)-4-Methyl-2-oxo-3-I (phenylacetyl) amino] l-azetldinesulfonic acid, potassium salt A) (3S)-3-Amino-4-methyl-2-oxo-l-azetidinesulfonic acid, tetrabutylanroonium salt (45-trans)-4-Methyl-2-oxo-3-[I (phenylmethoxy)carbonyl]amino]-1-azetidinesulfonic acid, potassium salt (352.4 ag; see Example 99} is dissolved in 20 ml of distilled water end treated with 373.5 mg (1 mmole) of tetrabutyl anroonium hydrogen sulfate. After 10 minutes stirring at room temperature, the solution is extracted three times with IQ ml portions of methylene chloride after saturation with sodium chloride. The methylene chloride is dried over sodium sulfate and evaporated in vacuo to give 536 mg of the tetrabutylammonium salt which is hydrogenated with 270 mg of 10« palladium on charcoal in 25 ml of dimethylformamide. The mixture is filtered through Celite and washed twice with 2.5 ml portions of dimethylformamide to yield the title compound in solution.

B) (3S-trans)-4-Methyl-2-oxo-3- Γ (phenylacetyl) amino]-1-azetidinesulfonic acid, potassium salt The crude (3S)-3-amino-4-methyl-2-oxo-lazetidinesulfonic acid, tetrabutylammonium salt from part A (the combined filtrate and washings) are treated at 0°C with 206 mg of dicyclohexylcarbodiimide, 153 mg of N-hydroxybenzotriazole, and 138 mg of phenylacetic acid. The reaction -137mixture is stirred at 0°C for 1 hour and then nt room temperature for 2 hours. The resulting precipitate is filtered, the filtrate is evaporated in vacuo, the residue is dissolved in 10 ml of acetone and filtered. The filtrate is treated with 25 ml of acetone saturated with potassium iodide and then 2Q0 ml of ether. The resulting solid (752.7 ag) is a mixture of the potassium and tetrabutylammonium salts of the title compound.

XO The solid is dissolved in 50 ml of 0.5 monobasic potassium phosphate and applied to an BP-20 column. Elution with water and then water-acetone gives several fractions that are combined and evaporated to give the purified tetrabutylammonium salt. An X5 agueous solution of this material is passed through Dowex 50W-X2 (K^fonn) to give the title potassium salt (121.4 mg). Trituration with acetone-hexane yields 104..6 mg of the title compound, melting point 211-213°C. 2q Analysis calc'd from C^jH^gNgOgSK-l/l HjO: C, 41.72; H, 4.09; N, 8.11; S, 9.28; X, 11.32 Founds C, 41.70; B, 4.01; N, 8.07; S, 9.01; X, 11.02 Example 101' (cis)-4-Methyl-2-oxo-3-I(phenylacetyl)amino]-1azetidinesulfonic acid, potassium salt A solution of 320 mg of (ΐ—-cis) -4-methyl2-oxo-3-[J (phenylmethoxy)carbonyl] amino]-l- azetidine30 sulfonic acid, potassium salt (see example 98) is prepared in 20 ml of water containing 483 mg of tetrabutylammonium hydrogen sulfate and adjusted to pH 5.5. 51333 -138Extraction with aix 25 ml portions of methylene chloride gives517.3 ag of oil. A solution of this material in 15 al of dimethylformamide is stirred with 400 ag of 10% palladium on charcoal in an atmosphere of hydrogen for 90 minutes.

The catalyst is filtered and the filtrate stirred with 150 ng of phenylacetic acid, 169 ag of N-hydroxybenzotriazole and 247 ag of dicyclohexylcarbodiimide for 7.5 hours. The solvent is removed in vacuo and the residue is dissolved in 2Q ml of acetone and filtered. The filtrate is treated with 25 al of 0.044 M potassium iodide in acetone. Dilution with an equal volume of ether gives e solid (330 ag} which is applied to a 50 ml HP-20 column in 20 ml of 0.05 M monobasic potassium phosphate. Elution with 200 ml of water followed by 1:9 acetone-water gives Rydon positive material in fractions (50 ml} 6-10. Evaporation of fractions 7-9 gives 81 mg of solid. Recrystallization from acetonitrile-water gives 46 mg of the title compound, which decomposes at >205°C. A second crop (6 mg) is obtained from the filtrate. A further 5 mg is obtained from fractions 6 and 10 by evaporation and recrystallization.

Analysis calc'd for C12813N2O5SX: C, 42.84; H, 3.89 N, 8.33; S, 9.53; X, 11.62 Pound: C, 42.75; H, 3.82; N, 8.32; S, 9.26 X, 11.63 51333 -139Example 102 J3S-f3a (Z) ,4a]]-3~H (2-Araino-4-thlazolyl) (methoxyimino) acetyl] amino] -4-methyl-2-oxo-l-azetidinesulfonic acid, potassium salt 5 A) (3S-trans)-4-Methyl-2-oxo-3- [I (phenylmethoxy) earbonyl Jamino]-1-azetidinesulfonic acid, tetrabutylammonium salt (35-trans) -4-Methyl-2-oxo-3-11 (phenylmethoxy) 10 carbonyl Jamino]-1-azetidinesulfonic acid, potassium salt (352.4 mg; see Example 9S) is dissolved in 20 ml of water and tetrabutylammonium hydrogen sulfate (373.5 mg) is added. The agueous solution is extracted three times with methylene chloride and the combined extracts are dried ever sodium sulfate. After removal of the solvent, 534.6 mg of the title compound is obtained.

B) [3S- [3o (Z) ,4S]]-3-I I(2-Amino-4-thiazoiyl) (methoxy20 imino) acetyl] amino] -4-methyl-2-oxo-l-azetidinesulfonic acid, potassium salt A solution of 534.6 mg of (3S-trans)-4-methyl2-OXO-3-II (phenylmethoxy) carbonyl] amino]-l-azetidinesulfonic acid, tetrabutylammonium salt in 20 ml of dimethylformamide is hydrogenated with 220 mg of 10% palladium on charcoal at atmospheric pressure for 2.75 hours; the uptake of hydrogen is 26.3 ml.

The mixture is filtered and washed twice with 2.5 ml of dimethylformamide. The filtrate and washings (total ca. 25 ml) are stirred under nitrogen with 161 mg of (Z)-a-(methoxyimino)-2-amino-4-thiazoleacetic acid, 136 mg of N-hydroxybenzotriazole 51333 -140 and 164.8 ag ot dicyclohexylcarbodiimide. The mixture ie stirred under nitrogen for about hours. The dimethylformamide is removed in vacuo and the gunmy residue is dissolved in acetone and filtered to remove urea. To the filtrate is added a solution containing 272 mg (0.8 mole) of perf luorobutanesulf onic acid, potassium salt in 0.8 ml of acetone. The slurry is diluted with an equal volume of ether and filtered to give 325.5 mg of crude product which is purified through chromatography on 75 ml of HP-20AG. Elution with 400 ml of water and 400 ml of (9:11 water: acetone mixture (50 ml fractions) gives 335 mg in fractions 3 to 10.

After trituration with acetone-hexane, 97.3 mg of an analytical sample is obtained from fractions 3-5. Similar trituration of fractions 6-1Q gives an additional 90.4 mg of product as a solid.

Analysis calc'd for C10H12N5OgS2K: C, 29.92; H, 3.01; N, 17.45; S, 15.97; X, 9.74 Found: C, 30.32; H, 3.49; N, 15.82; S, 13.95; X, 10.45 NMR(D20, 1.57 (3H, d,j»7), 3.97 (3H.S), 4.3O(1H, d of q, j-7.3), 4.7O(1H, d. .,-7), 6.95ppm(lH,S).

Example 103 I3S-I3a (Z) ,4811-3-1 [ (2-Amino-4-thiazolyl) Hl-carboxy1-methyl ethoxy) imino] acetyl] amino] -4-methyl-2-oxol-azetidinesulfonic acid, dipotassium salt A) N-Benzyloxy-t-boc-threonine amide A solution of 8.76 g of t-boc-threonine and the free amine from 6.4 g of O-benzylhydroxylamine HCl (ethyl acetate-sodium bicarbonate liberation) in 100 ml of tetrahydrofuran is treated with 6.12 g 513 9 3 -141of N-hydroxybenzotriazole and 8.24 g of dicyclohexylcarbodiimide in 2Q ml of tetrahydrofuran.

The mixture is stirred under nitrogen for 26 hours, filtered, and evaporated in vacuo.

The residue is chromatographed on a 300 g silica gel column (elution with chloroform and chloroformethyl acetate (3:11) yielding 7.2 g of compound. Crystallization frcm ether-hexane gives 4.18 g of the title compound.

S) (3S-trans)-N-8easyloxy-3-^-butoxycarbonylamino4-methylazetidinone A solution of 12.67 g of N-benzyloxy-t-boethreoniae amide, 11.5 g of triphenylphosphine, and 6.23 ml of diethylazodicarboxylate in 380 ml of tetrahydrofuran is stirred under nitrogen for shout 16 hours. The solution is evaporated and chromatographed on a 900 gram silica gel column. Elution with chloroform-ethyl acetate (3:1) gives 13.69 g of compound that crystallizes from ether-hexane to yield 9.18 g of the title compound.

C) (3S"traasi)"3<"t"Butoxycarbonylamino-l-hydroxy25 4-methylazetidinone A solution of 9.18 g of (3S-trans)-N-benzyloxy3-t-butoxycarbonylamino-4-methylazetidinone in 300 ml of 95S ethanol is stirred in an atmosphere of hydrogen with 1.85 g of 10% palladium on charcoal.

After 141 minutes the slurry is filtered and evaporated in vacuo. The residue is recrystallized from ether-hexane to yield 5.12 g of the title compound -142D) (35-trans)-3-t-Butoxycarbonylamino-4-methylazetidinone A eolution of 4.98 of (3S-trans)-3-t-butoxycaxbonylaraino-l-hydroxy-4-methylazetidinone in 20Q ml of methanol is treated with 132 ml of 4.5 M ammonium acetate and then 66 ml of 1.5 M titanium trichloride and stirred for 4.5 hours. The aqueous solution is diluted with an equal volume of 8% sodium chloride and extracted with ethyl acetate to give 10 3.48 g of crude product. Reerystallization from ether-hexane yields 3.3 g of the title compound.

E) (35-trang.l-3-Benzyloxycarbonylamino-4-fflethylazetidinone a solution of 3.3 g of (3S-tran>)-3-t-butoxycarbonylamino-4-methylazetidinone in 10 ml each of dichloromethane and anisole is cooled to 0°C and 112 ml of trifluoroacetic acid is added. The solution is stirred for 90 minutes and evaporated in vacuo (benzene added and evaporated three times). The residue is dissolved in 70 ml of acetone and the solution is adjusted to pH 7 with 5t sodium bicarbonate solution. A total of 5.33 g of benzyl chloroformate is added over 1 hour at pH 6.5-7.5.

The mixture is stirred for 30 minutes at pH 7, diluted with 100.ml of saturated salt, and extracted with ethyl acetate (three 400 ml portions). The residue obtained by evaporation is chromatographed on a 1 liter silica gel column. Elution with chloroform-ethyl acetate (4:1) gives 2.19 g of compound. Crystallization from ether-hexane yields 1.125 g of the title compound. -143F) (3S°trans)-4-Methyl-2-oxo-3-[ ( (phenylmethoxy)earbonyl]amino]-1-azetidinesulfonic acid, tetrabutylammonitaa salt A solution of 600 mg of (35-trans)-35 benzyloxycarbonylamiao-4-methylazetidinone in ml of dimethylformamide is cooled to 0°C and 4 ml of 0.8 M sulfur trioxide in dimethylformamide is added. The solution is stirred at room temperature under nitrogen for 1 hour and poured into 80 ml of cold 0.5 K monobasic potassium phosphate (adjusted to pB 5.5). The solution is extracted with three 50 ml portions of methylene chloride (discarded) and 863 mg of tetrabutylaraaonium bisulfate is added. She resulting solution is extracted with four 75 al portions of methylene chloride. The combined organic layer is washed with 8% aqueous sodium chloride, dried, and evaporated in vacuo yielding 1.54 g of the title compound.

G! [35-[3g (2) ,4e]]-3-iI(2-Amino-4-thia2olyl)[ (l-diphenylmethoxycarbonyl-l-methylethoxy) imino] acetyl] amino] -4-methyl-2-oxo-l-azetidinesulfonic acid, potassium salt A solution of 1.54 g of (3S-trans)-4-methyl2-oxo- 3-11 (phenylmethoxy) carbonyl] amino] -1-azetidinesulfonic acid, tetrabutylammonium salt in 45 ml of dimethylformamide is stirred in an atmosphere of hydrogen with 800 mg of 10S palladium on charcoal for 2 hours. The catalyst is filtered and the filtrate stirred for about 16 hours with 51333 -1441.24 g of (Z)-2-amino-a-f (1-diphenylmethoxycarbonyl-lmethylethoxy) imino]-4-thiazoleacetic acid, 0.4 g of N-hydroxybenzotriazole, and 580 mg of dicyclohexylcarbodiimide. The slurry is evaporated in vacuo and the residue is triturated with 20 ml of acetone and filtered. The filtrate (plus 2 ml of washings] is treated with 868 mg of potassium perfluorobutanesulfonate in 3 ml of acetone. Dilution with 75 al of ether gives a solid that is isolated by decantation of the mother liquor, trituration with ether, and filtration to give 0.91 g of the title oompound. The mother liquor is diluted with a further 1Q0 al of ether to give a second crop, 0.45 g, of the title compound.

H) 135-[3a(Ζ),4 B]]-3-11 (2-Amino-4-thiazolyl)I (Ιο arb oxy-l-methyl ethoxy) imino] -acetyl] amino] -4methyl-2-oxo-l-azetidinesulfonic acid, dipotassium salt A slurry of 140 mg of [3S-[3α(Z),4B]]-3[ [ {2-amino-4-thia zolyl) [ (1-diphenylmethoxycarbony11-methylethoxy) imino] acetyl ] amino] -4 -me thy 1-2oxo-l-azetidinesulfonic acid, potassium salt (first crop) in 0.5 al of anisole is stirred at -12°C under nitrogen and 2.5 al of cold (-10°C) trifluoroacetic acid is added. After 10 minutes, 10 ml of ether and 5 ml of hexane are added and the resulting slurry is stirred for 5 minutes at -12°C, and allowed to warm to room temperature. The solid is isolated by centrifugation and washed twice with ether. A solution of this solid in 5 ml of cold water is SX333 -145imaediately adjusted to pH 5.5 with 0.4 N potassium hydroxide and then applied to an 80 ml HP-20AG column. Elution with water gives 72 mg of the title compound in fractions (10 ml) 7-11 after evaporation (acetonitrile added and evaporated three times) and trituration with ether, m.p.~250°(dec.). Analysis calc'd for C^Bj^NjOgSjXj: C, 30.51; H, 2.95; N, 13.69; S, 12.53; X, 15.28 Found: C, 29.63; H, 3.20; H, 12.96; S, 11.94; X, 12.78 NMR(D2o) 1.46(5, 6H), 1.58(1H, d, j«7) 4.28(lH,'d of q, j"7, 2.5), 4.67 (IH, d, j*2), 6.95ppm(S, IH).

The remaining 1.22 g of I3S-I3e(Ζ1,4β]J-315 [I (2-amino-4-thiazolyl) 1(1-diphenylmethoxycarbonyl1-methylethoxy) imino] acetyl] amino-4-methyl-2oxo-1-azetidinesulfonic acid, potassium salt (crops 1 and 2) are treated as above (4.2 ml anisole, ml of trifluoroacetic acid, 13 minutes at 2Q -15°C). Chromatography on a 300 ml HP-20AG column gives 694 mg of the title compound in fractions (60 ml) 6-9 after treatment as above.

Examples 104-133 Following the procedure of Example 11, but substituting the acid listed in column X for (Z) -2-amino-a- [ I Ihydroxy (phenylmethoxy) phosphinyl] methoxy]imino]-4-thiazoleacetic acid, yields the compound listed in column XX. ι ο 1 · Ο JJ Η S&8 J μ » itil Μϋ π & · 9 ·οΐ j* a —j·* ‘Pfiifc0* 75838 •to >» · 8»<μ μ5*? *♦» si*3?o *-·β «5 · & ** C Α» Ό ·. a Μ « «Η ο Μ ο ϋ · 1 < ό Ο Γ Ο Α» I to Β Ο ·. 2il*f « Ο»«ιίί to to » I φ >» ϋυ η ι *J ttO ι η · «η « · ο οβ «» <Μ 4 «Η «Η t A^S— >, 2 *4 c aim » β *ι 5 "" Β & . = 83ο 8| 7 5 S3’1 7 ?·£ s J =7 85°.» « -π X β β» 857 Μ Μ Ό <Η < © 3 2* 33 «ο ?8 Η I V *» ·*< S8 is Al · • W »4 ο £3 1? «4 Γ3· Η* 8 —» -Η I β η ·η 7? ?s. 8ϊ r ? W Cf TJ I *H Α ϋ to * x c o •H *H N Al tt · u u xs *H C ¥•8 to C ii Cl I I w e4 I ? ; 8 ** T & Pi ~ u 5 8 tt tt X? ?: H «Η I Al s| n a * *u I Cf Η I ΐ7 Al C Π T c PS ** u tl 8 to 0 C Al •H ft Ν V tt tt U ft XS Ή to O.TJ Λ 3 u 55 3 & A? £8 •5 • ba U t · « e 8S «η e Π • tt e o • * -4 «□ X £ A A AJ I Π il i & 8* 7" &Ό C< --4 w4 — HO "2 * I · 0 a a*t I to At — a · oc ι □ -147I 9 ri c q £3 2. fie» Tec rt a «» ι ri *— rt rt :i 2 >» X rt 0 * I ri H tt rt a ι a rt a S s 8 * a® 8=8 rt > ri St! & o Αντί 8 ώ 7 4 ο <81 . ί 2 rt U ϊ tol Ο Ο · C rt vl a e χ Ο * &<μ ο * rtO ** 9 η rt Μ 10 > β rt rt C 0 rt ri η ·α c rt rt rt ri O £ · fit •?r I rt rt Si ri rt 8 B " 0 β = 8S I rt TSS n 5 Ρ υ ** δ **ο CO X rt © « Α» 9 Ο — β rt N > — Αί β oj β ri 28 8. ri to a o • I to C— to β e o rt «1 rt c c >rt O · x « *-u ri 3 rtO Ϊ- 9 tt rt rt C Α Ό C 9 W rt rt x a a* o 6b U 9 B· H K rt · c U fi rt rt v rt I ri >» V O rt £ rt X C ι <2 o i ν o T ** M rt w -BID •rt Vsrtrt I rt Q · 233 = A 5 S ~ | ^rt Μ η H ι κ >« a rt p ri a ι I 9 ri ϋ 0 — ι a cu « rt» rt 08 5 >· · ** fi c « ca rt · rt a w C rt a > § ?s rt to rt a ι to rt ι ri 0 a X c to X a I I c e e a rt 13, X 0 to owe E to « rt I 0 tt N rt. I G — 0 C I K to non — I s = 73 8*0*0 ι e e >» rt I e >»rt ri rt e 9 e v p a rt rt i β a S.S3 & Q. U V 5» I rt»rt C tt rt ri rt **>,·*, •—Bert I rt β β «π v I I I rt rt O rt I Κ O ~ ri ? 8 8*3 N 85rt =31s m | rt rt " rt > a ι ι ri a rt«m · a i o to ri In*. tt «-» a V O ι β λ ·η a ~ c α. o vl a a ι a ♦ rt »» * OS >» O to U ** Λ C rtO ca ri rt s O **» a a o rt h-. 2 3 £ cb ι rt V I rt *o a c rt U -» a a to e a a a rt C ►. a X N ri c o S 5 m rt< b ι a tt « ι rt X > o e I rt ee »0 — 0 I M 8 a I V A rt OS E I 0 ri re C rt I rt « :s· 5z5 £ ϊ ι a a rt u a a ai "*o 8» c >»rt · η a v 0 rt rt · M p- U Ul a > a a| rt κ v χ o to ri £ rt » l ϋ CO tt a oo I «u m 8>rt rt K 9 rt rt o a ι β x a o a ri c rt I a rtrt tt fi V , rt rt rt ri *rt >AJ fi ** c a rt I a a 0 η χ a & 1 β. ι F-»rtH 0» ** I I c ta c· ρ ·η ** | fi A* S S ?*3 " 0 tt fi ι to η a >» C to a rt x ri c. a — o ι a cm a I rt 0 0 X N 7£ rt I &Σ =1 ι *5 β X I 0 gf E * a s ι x tt o £8 o -148u t u ο ι a -8 -78 ί7ΐ 0** M Q 0 « C Λ» * si i © «0 51 I X© 2U O · 0 U p38 φ: Ξ23.5 α>:& -h? *H83 β Μ X© © *-***»*» a e · © — 8,31 .8 , S’ I ** o » >8 ·οϋ S32S «8 ** © Μ © ** > © β ·— V 9 © Ss s*· X·— © » β Ο *0 G • β © © X © © Λ» — 5 S3 73 r1 © X © m I C I © ski ϊ>5| ί * ·* -1 3 8. η I © ri 0 3 © Λ» 8 ·, ri ο 3™ 0 · "ο° © ο ο X© © 3δΜ* 4ί X 9© ή 3 & — 6 _ ? © © © I Λ» © ri 9 © 338 • υ . ’’Ό I © © © ο © I X 0 © © C I I 0 ο © © X © © ?£§ ξ 8 3 e © 5 5 8.

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Μ Τ3 I · Μ Μ Ν Α > 4 Κ It! « Q Μ I I 4 f4 C* · > ι Ή 5 ο Ηϊ« sis ο ι β π Λ Ο · «Μ ι £«1 *» Μ«4 Ο Β · υπ «~3 «Ν TJ Μ U Ο Μ I W I Μ > C I -153Example 134-135 Following the procedure of Example 70, but substituting the compound listed in column I for (3S(Z) J-3-[[(2-amino-4-thiazolyl,((2-(diphenylmethoxy, -2-oxoethoxy] imino) acetyl] amino] -2-oxo5 1-azetidinesulfonic acid, potassium salt, yields the compound listed in column II. -154513S3 Coluwi I Coluwi ix I g> m s a e I 0 4 A F* A M A M A 4 M m 3 M I — A I 4 AU2 "?5 8 Μ Μ A β S I · -* M f M gars 5 5? 3 518. 45? 3 ι · n · |Ιλ: A i o a ?··μ N S 4 » — Κ·—Ό ·— Ο Μ -H "£ >, O I A A 4 <*> · · I W O O ~ S 4 M e Ν A Ο O OS 1 Μ M m A B □ — Ό* a I fr U 5 . c A F-e 0 • mm 5 &9 • fa Β M £ 4 μ fM ,2,3 · "77 a *h " S · Q* I c Μ Ϊ η m 4 4 ι « ι x 85? ?878 a«|s =3is " I M 4 I m s a η ι 4J __ I r+« I 0Q9 „ c S o «μ a IBS· !M 4 C 4 I f-w · A z1£& Sc § 7 * " £ c e W Μ Μ M m 4 Β o — 044 Si 393 -155Example 136 [3a (2) ,4a)-3-[[(2-Amino-4-thiazolyl)(methoxyimino) acetyl]amino]-4-methyl-2-oxo-l-azetidinesulfonic acid, potassium salt A solution of 51.8 mg of (cis)-4-methyl5 2-oxo-3-[[(phenylmethoxy)carbonyl)amino]-1azetidinesulfonic acid, potassium salt and 51 mg of tetra-n-butyl-ammonium biaulfate in 5 ml of water is extracted with methylene chloride (four 10 ml portions) to give 81 mg of oil.

This is stirred in an atmosphere of hydrogen for 2 hours with 40 mg of 10* palladium on charcoal in 4 ml of dimethylformamide. The catalyst is filtered and washed with 1 ml of dimethylformamide. The filtrate and washings are combined and stirred for about 16 hours with 31 mg of (Z)-2-amino-a-(methoxyimino)-4thiazoleacetic acid, 27 mg of N-hydroxybenzotriazole, and 31.5 mg of dicyclohexylcarbodiimide. The solution is evaporated in vacuo and the residue triturated with 3 ml of acetone. The resulting slurry is centrifuged and the liquid treated with 51 mg of potassium perfluorobutanesulfonate. Dilution with 5 ml of ether and filtration gives a solid. Chroma25 tography on HP-20 AG (40 ml) gives Rydon positive material in fractions (20 ml) 3-5 (elution with water). Evaporation and ether trituration gives 23 mg of product, as a hygroscopic solid. Analysis calc'd for ci0H12N5°6S2K: C,29.91; H.3.01; N,17.44 Found: C,29.30; H,3.31; H,16.66 NMR(D2O5 1’40(3h' 2=7), 3.97 (3H,S), 4.46 (IH, apparent pentet, ;»7), 5.37(IH, d, j-7), 6.97 ppm(lH,S). -156gxample 137 (3S-cis)-3-Amino-4 -methyl-2-oxo-l-aretldinesulfonic acid A) t-Boc-l-allothreonine A suspension of 6.72 g ot 1-allothreonine ia 70 al of 50« aqueous dioxane is treated with 9.45 al of triethylamine and 18.1 g ot t-butylpyrocarbonate. The resulting aixture is stirred at room temperature for 4 hours, and then diluted with 70 al of water and 140 al of ethyl acetate. After thorough shaking, the layers are separated and the organic layer ia washed with 30 al of 2:1 water:brine. Combined aqueous layers are then back-extracted with 70 ml of ethyl acetate. The aqueous layer is cooled in an ice bath and 10« potassium bisulfite solution is added to pH 2.3. The acidified solution is extracted with ethyl acetate (four 150 ml portions). Combined organic layers are dried over anhydrous sodium sulfate and stripped of eolvent to give 9.13 g of the title compound. 8) N-Methoxy-t-boc-l-allothreonine amide t-Boc-l-allothreonine (9.13 g) is dissolved in 85 ml of water and 41 ml of IN potassium hydroxide solution. Methoxyamine hydrochloride (5.22 g) and 8.67 g of l-ethyl-3,3-(dimethylaminopropyl) earbodiimide *BC1 are added. The 51353 -157mixture ie stirred at room temperature for 4 hours and then saturated with sodium potassium tartarate. The resulting mixture is extracted with ethyl acetate (four 150 ml portions) and the organic layer is dried over anhydrous sodium sulfate and stripped of solvent to give 7.38 g of the title compound as a solid.

C) O-Methanesulfonyl-N-methoxy-t-boc-l—allothreonine 10 amide N-Methoxy-t-boc-l-allothreonine amide (7.32 g) is dissolved in 4Q ml of pyridine and cooled to -20°C under nitrogen. Methanesulfonyl chloride (3 ml) is added dropwise by syringe over a -minute period. The resulting mixture is slowly warmed to 0°C and stirred at that temperature for 3 hours. Ethyl acetate (500 ml) is added and the solution washed with 250 ml of ice-cold 3N SCI solution, then 100 ml of 5% NaHCO^ solution. 2Q The ethyl acetate layer was dried over anhydrous sodium sulfate and stripped of solvent to give 8.64 g of the title compound as a white solid.

D) (3S-cis)-3-t-Butoxycarbonylamino-l-methoxy25 4-methylazetidinone Ο-Methanesulfony1-N-methoxy-t-boc-1-aliothreonine amide (8.64 g) is dissolved in 530 ml of acetone and 11 g of solid potassium carbonate is added. The mixture is slowly heated to 65 C under nitrogen and stirred at this temperature for one hour. The reaction mixture is then -158filtered through Celite and the filter cake is washed with ethyl acetate. The filtrate is concentrated and the residue is taken up in 250 al of ethyl acetate. The ethyl acetate solution is washed with 100 si of IN hydrochloric acid solution and 100 al of 5* sodiua bicarbonate solution. The ethyl acetate layer is dried over anhydrous sodiua sulfate and stripped of solvent to give 6.63 g of crude product.

B) (3 S-cis) —3—t-Butoxyc arbonylamino—4 -methylazetidinone Sodium (1.35 gl is dissolved in about 300 ml of liquid ammonia at -50°C and 5.87-g of (38-cis)3-t-Butoxyearbonylamino-l-methoxy-4-methylazetidinone in 35 ml tetrahydrofuran is added dropwise via syringe. An additional 10 ml of tetrahydrofuran is used for rinsing. Near the end of the addition about 100 mg of additional sodium is added. The mixture is stirred for five more minutes, then quenched by adding 3.35 g of solid ammonium chloride in one portion. Ammonia is blown off with a nitrogen stream and 250 ml of ethyl acetate is added to the residue. After filtration and washing the solid with ethyl acetate, the combined filtrate is stripped of solvent to give 4.82 g of the title compound. -159P) (35-cis)-3-t-Butoxycarbonylamino-4-methyl2-oxo-l-azetidlnesulfonic acid, tetrabutyl ammonium salt (3S,4R]-3-t-Butoxycarbonylamino-4 -methylazetidinone (4.98 gl is dissolved in 30 al of dimethylformamide. Pyridine-sulfur trioxide complex 11.9 g is added and the mixture is stirred at room temperature under nitrogen. After 14 hours stirring, an additional 1.8 g of pyridine-sulfur trioxide complex is added and stirring is continued for 80 hours. The reaction mixture is poured into 700 ml of 0.5 M monobasic potassium phosphate solution and washed with methylene chloride (three 300 ml portions). tetra-n-Butylammonium bisulfate (8.45 g) is added to the agueous solution and the mixture is extracted with methylene chloride (four 300 ml portions). The combined methylene chloride layers are dried over anhydrous sodium sulfate and stripped of solvent to give 10.76 g of the title compound as a gum.

G) (3s-cis)-3-Amino-4-methyl-2-oxo-l-agetidinesulfonic acid (35-eis) - 3-t-Butoxyc arbonylamino-4 -methy12-oxo-l-azetidinesulfonic acid, tetrabutyl ammonium salt (10.76 g) is dissolved in 50 ml of 95-97% formic acid and stirred for 4 hours under nitrogen. A small amount of product from a previous reaction is added as seed and the mixture is stirred for one more hour. The 51333 -160mixture is stored in the freezer for about 16 hour* and the frozen mixture is warmed to room temperature and stirred for en additional hour. The solid formed ia filtered and washed with methylene S chloride to yield 982 ag of the title compound.

The filtrate is diluted with 1 liter of methylene chloride and kept at -20°C for 4 hours. The precipitate that forma is recrystallized from water-methanol-acetone to give an additional 167 ag of title compound.

NMR(D20) 1.63.(38, d, J-6.5 cps), lR(nujol)1775 cm-1.

Example 138 13S- I3a ( Z), 4aJ ] -3-1 H-Amlno-4-thiazolyl) methoxyimino) acetyl] amino-4-methyl-2-oxo-lazetidinesulfonic acid, potassium salt 15 A solution of 201 mg of (Z) -2-aaino-e(methoxyimino)-4-thiazoleacetic acid and 153 mg of N-hydroxy-benzotriazole monohydrate in 3 ml of dimethylformamide is treated with 206 mg of dicyclohexylcarbodiimide. The mixture is stirred at room temperature for 20 minutes under nitrogen and a solution of 180 mg of (3S-cis)3-amino-4-aethyl-2-oxo-l-azetidinesulfonic acid and 0.14 ml of triethylamine in 2 ml of dimethylformamide is added (an additional 1 ml of dimethylformamide is used for rinsing) and the mixture is stirred for about 16 hours. The slurry is evaporated in vacuo, triturated with 12 ml of acetone, centrifuged and the liquid treated with 338 mg of potassium perfluorobutane3Q sulfonate. Dilution with 10 ml of ether and filtration gives a solid product which is chromatographed on 200 ml of HP-20 resin eluting with -161water. Fractions (20 ml each) 18-30 are combined and lyophilized to give 274 mg of the title compound as a hygroscopic solid.

Anal, calc'd. for c10Hi2N5O6S2K: C,29.91; H, 3.01; N.17.44 Found: C.30.03; H,3.21; N.17.06 NMR(D20) 1.40 (3H, d, j—6.5), 3.98(3H,S), 4.48 IH, d of t, ,=6.4, 5.5), 5.36(1H, d, ,=5.5) 6.97 (1H,S).

Example 139 (3S-fcrans)-3-Amino-4-methyl-2-oxo-l-azetidine-sulfonic acid A) Threonine, methyl ester, hydrochloride Dnder an atmosphere of nitrogen, a flask containing 500 ml of methanol is cooled to -5’C (ice/brine) and 130 ml (excess) of thionyl chloride is added at such a rate as to maintain the reaction temperature between 0° and 10°C. After recooling to -5°C, 59.5 g of 1threonine is added and the mixture is allowed to reach room temperature and stirred for 16 hours. The mixture is concentrated and evacuated at 10-1 torr for 2 hours to yield a viscous oil. This material is used directly in the following step. -1628) Threonine amid· Tha crude product from part A ia dissolved in 2.5 1 of methanol and cooled to -5°C (ice/brine). The solution is saturated with ammonia gas, the cooling bath is removed end the sealed vessel is allowed to stand for 3 days. After removing the bulk of the unreaeted ammonia via aspirator, 100 g ot sodium bicarbonate and 50 al of water is added and the mixture is stripped to a viscous oil.

Cl Benzyloxycarbonyl threonine amide The crude product from part B (already containing the requisite amount of sodium bicarbonate is diluted to a volume of 1 liter with water. To this rapidly stirring solution, g (88 ml of 90% pure material) of benzyloxycarbonyl chloride is added as a solution in 80 ml of tetrahydrofuran over a 1 hour period. The reaction mixture is then stirred for an additional 16 hours and extracted with ethyl acetate (one 500 al portion, two 250 ml portions). The combined extracts are dried over magnesium sulfate and concentrated. The crystalline residue is then dissolved in 250 ml of hot ethyl acetate and 300 ml of hexane is added followed by boiling until a clear solution is reached. Cooling and filtration of the crystalline mass give, after drying, 104 g of the title compound. 51303 —163— D) Benzyloxycarbonylthreonine amide, O-mesylate Under an atmosphere ot argon, 100 g of benzyloxycarbonylthreonine amide ia dissolved in 400 ml of anhydrous pyridine and cooled in an ice/salt bath. To this stirring solution, 36.8 ml (54.5 g) of methanesulfonyl chloride is added over a 15 minute period. After 2 hours of stirring an additional 0.3 equivalents of methanesulfonyl chloride is added. The reaction is then stirred for 1 hour and poured into a mixture of 1.5 1 of ice and 2 1 of water. The resulting slurry is stirred for about 30 minutes and filtered. Drying of the crude product at 60°C for about 16 hours in a vacuum oven gives 109 g of the title compound.

E) N-Sulfonyl benzyloxycarbonylthreonine amide.

O-mesylate, tetrabutylammonium salt A solution of 2-picoline (17.8 ml) in 2Q 90 ml of methylene chloride is cooled to -5°C (ice-brine) and chlorosulfonic acid 5.97 ml is added at such a rate as to maintain the internal reaction temperature below 5°C. The resulting solution is added via canula, to a suspension of 7.56 g of benzyloxycarbonylthreonine amide, O-mesylate in 120 ml of methylene chloride. The resulting heterogeneous mixture is refluxed for about 16 hours yielding a clear solution.

The solution is poured into 500 ml of pH 4.5 30 phosphate buffer (0.5 M) and further diluted with 120 ml of methylene chloride. The separated 51333 -164organic layer la then washed once with 100 ml of buffer solution and the combined agueous phases are treated with 10.2 g of tetra-n-butylannonium hydrogensulfate and extracted with methylene chloride tone 300 ml portion and two 150 ml portions). After drying the combined organic extracts over sodium sulfate, the solution ia concentrated to yield 12.7 g of a foam.

Fl (3S-trans) -3-Amino-4-methy 1- 2-oxo—1—azetidinasulfonic acid A mixture consisting of 5.52 g of potassium carbonate in 20 ml of water and 160 ml of 1,2diehloroethane is brought to reflux and 15.5 mole of Ν-sulfonyl benzyloxycarbonylthreonine amide, O-mesylate, tetrabutyl ammonium salt is added in 20 ml of 1,2-dichloroethane (20 ml used as a rinse). After refluxing for 30 minutes, the mixture is poured into a separatory funnel, diluted with 50 ml of water and 100 ml of methylene chloride and the phases split. The resulting organic phase is dried over sodium sulfate and concentrated to yield crude (3S-trans)3-benzyloxycazbonylamino-4-methyl-2-oxo-l-azetidinesulfonic acid, tetrabutylammonium salt. The crude azetidinone is treated in 250 ml of ethanol with 0.8 g of 5% palladium on charcoal catalyst and hydrogen is bubbled through the solution.

After 90 minutes the mixture is filtered through Celite with 50 ml of ethanol used as a rinse. 51333 -165The addition of 1.2 ml of formic acid to this solution causes an immediate precipitation of the title zwitterion which is filtered after stirring for 1 hour to yield, after drying at torr for 1 hour, 1.1 g of product. A second crop of product is obtained upon concentration of the filtrate and addition of more formic acid to give 1.3 g of the title zwitterion. m.p. > 218°dec., [α]β " -41.1(01, HjO).

NMR(D20) 1.58(3H, d j«7) , 4.80(2H,M).

Examples 140-143 Following the procedure of Example 138, but substituting (3S-trans)-3-amino-4-methyl-2-oxo15 1-azetidinesulfonic acid for (3S-cis)-3-amino-4methyl-2-oxo-l-azetidinesulfonic acid and the acid listed in column I for (Z)-2-amino-a-(methoxyimino)4-thiazoleacetic acid, yields the compound listed in column II. 513S3 52Λο 338* 33ο ό © Ό I © 0 X’S© 8 A Τ* ϊ ου *- Η 0 *0 = i 8 S3 = S3.S AA 5 2 c ' '<8* I Έ8 c ΐ 3 ί ε = •h >1 © ο li && © Ν Ό Ό 9 © © «Μ © Q I Ο 0 © Ν ** 0 U " Ό © I © © © Β 0 2.3 © ο Οι 0 I 2 ϊ © 0 » Ν © 6 Α2 έ*ο © e 0 © I Β ο Μ © £ © I ·—t -4 I I β ο £° 5Α2 ri U Η8 I © υ · I ο © © I © ri 0 as e © ο S3 ** 0 Μ Ο I 0 Ο 0 I C Γ.2 ** ο» 513 9 3 -167Example 144 [35-f3a(Ζ),4Β)]-3-[[(2-Amino-4-thiazolyl)[[1,1dimethyl-2-[(4-nltrophenyl)methoxy}-2-oxoethoxy)lmlno)acetyl)amino]-4-methyl-2-oxo-l-azetidine5 gulfonic acid, potassium salt To a elurry of (35-trans)-3-amino-4-aethyl2-oxo-l-azetidinesulfonic acid (0.36 g; see example 139) in dry dimethylformamide [30 ml) under nitrogen at 26°C is added triethylO amine (309 μΐ). After about 5 minutes a clear solution is obtained and tf 1-2-amino-a-[11,1dimethyl-2-1 (4 -nitrophenyl) me thoxy] -2-oxoethoxy] imino]-4-thiazoleacetic acid (0.816'g) is added followed by N-hydroxybenzotriazole (0.334 g) and dicyclohexylcarbodiimide (0.453 g). The mixture is stirred for twelve hours at 26°C, whereupon the solvent is removed in vacuo, and the residue is triturated with acetone (30 ml). After stirring five minutes, the solids are’ θ removed and the filtrate is treated with potassium perfluorobutane sulfonate (3.680 g) in acetone, (5 ml). Addition of ether (approximately 40 ml) affords a precipitate which is collected and dried in vacuo (1.073 g; second crop 0.066 g; total of 1.14 g).

Anal, calc'd. for C^H^NgO^SjX·! H2O: C, 38.33; H, 3.70; N, 13.41; S, 10.23; K, 6.24 Found: C, 38.30; H, 3.63; N, 13.41; S, 9.88; K, 5.98 51383 -168gxample 145 (3a(Z),4a)-3-[ [ (2-Amino-4-thlazolyl)[(1-carboxyl-methylethoxy ) imino] acetyl ] ani no] -4-methyl-2oxo-1-azetidinesulfonic acid, potassium «alt (1:2) A) (3a(Z),4a]-3-[[2-Amino-4-thiazolyl)I(1diphenylmethoxycarbonyl-l-aethylethoxy) imino]ecetyl 1 amino] -4-methyl-2-oxo-l-azetidinesulfonic acid, potassium salt A solu'-xon of (cis)-4-methyl-2-oxo-3-[ [ (phenylmethoxy) carbonyl] amino] -1-azetidinesulfonic acid, tetrabutylammonium salt (201 mg.; prepared from the corresponding potassium salt of example 98 as described in example 136) in 5 al of dimethylformamide is stirred with 90 mg of 10% palladium on calcium carbonate in an atmosphere of hydrogen for 2 hours. The slurry is filtered and the filtrate is stirred for about 16 hours with 146 mg of (Z) -2-anino-a- [l-diphenylmethaxycaiixjnyl-l-inethylethQjQf) imino] 4-thiazoleacetic acid, 73 mg of dicyclohexyleaibodiimid and 51 mg of N-hydroxybenzotriazole under nitrogen. The slurry is evaporated in vacuo and triturated with 4 ml of acetone. The slurry is filtered and the solid washed twice with 2 ml portions acetone. The filtrate and washings are combined and treated with 113 mg of potassium perfluorobutanesulfonate. Dilution with 24 ml of ether gives a solid that is isolated by centrifugation and washed three times with ether yielding 186 mg of the title compound. 1 ’·» '' 3 -169B) [3o(Z) ,4o)-3-[ I (2-Amino-4-thiazolyl) f 1-carboxy— 1-methylethoxy) iaino) acetyl lamino)-2-methyl-4-oxol-azetidinesulfonic acid, potassium salt (1:2) A Blurry of 186 mg of [3α(Z),4a]-3-1 [2-amino4-thiazolyl) [ (1-diphenylmethoxycarbonyl-lmethylethoxy) imino) acetyl] amino] -4-methyl-2-oxol-azetidinesulfonic acid, potassium salt in 0.6 ml of distilled anisole is cooled to -12°C and 3.0 ml of distilled trifluoroacetic acid (at =10°C) is added. The solution is stirred for 10 minutes and 12 ml of ether followed by 6 ml of hexane are added. After 5 minutes at =10°C and stirring for 15 minutes at ambient temperature, the solid is isolated by centrifugation and washed four times with ether to give 141 mg of material. This is dried in vacuo, powdered, dissolved in 5 ml of cold water and immediately adjusted to pH 5.6 with 0.4 N potassium hydroxide. The solution is applied to a 100 ml HP-20AG column and eluted with water. Fractions (10 ml) 0-12 are combined and evaporated in vacuo (acetonitrile is added three times and evaporated. The residue is triturated with ether to give 101.7 mg of product, as a hygroscopic solid.

Anal. Calc'd for εΐ3Ν15Ν5Ο8ε2,2Κ! C, 30.51; H, 2.95 N, 13.69; S, 12.53; X, 15.28 Found: C, 30.11; H, 3.26; N, 13.35; S, 12.12; K, 15.02 51333 -170gxample 146 [3S-(3a (8),48] 1-3-Π (2-Aalno-4-thlazolyl)-((1carboxy-1— aethylethoxy) ini no] acetyl lamino] -4methyl-2-oxo-l-azetidinesulfonic acid I3S-I3a(X),4 β1]-3-1 ((2-Amino-4-thiazolyl)[ (1-carboxy-l-methylethoxy) iaino] acetyl lamino] 4-methyl-2-oxo-l-azetidinesulfonic acid, dipotassium salt (87.3 mg; see example 103) is dissolved in 1.38 ml of water, cooled to 0°C, treated with 0.34 al of IH hydrochloric acid and the resulting crystals separated by centrifugation. The wet solid is dissolved in methanol, filtered, concentrated to about 0.5 al and mixed with 1 ml of water, giving 55.9 mg of the title compound.

Example 147 (3S-(3a(Z),4B]]-3-(I(2-Amino-4-thiazolyl)[(lcarboxy-l-methylethoxy)imino]acetyllanlno3-A-methyl-2oxo-1-azetidinesulfonic acid, sodiun salt λ 99.7 mg sample of I3S-I3a(Z),48))-3[[ (2-amino-4-thiazolyl) [(1-carboxy-l-methyle thoxy) iaino] acetyl] amino] -4-methy 1-2-oxo-laz.etidinesulfonic acid is mixed with 0.207 al of IN sodium hydroxide and the resulting mixture is gently warmed to dissolve the remaining solid, water is removed azeotropieally with acetonitrile and the residue is crystallized from a mixture of 0.5 ml of methanol (to dissolve the residue) and 1 ml of acetonitrile, giving 8Ϊ.8 mg of solid, λ second recrystallization from 0.8 ml of methanol gives 47.9 mg, a third from 0.24 ml of methanol and 0.24 al of absolute ethanol gives 44.8 ag, and -171a fourth from 0.225 ml of methanol and 0.225 ml of absolute ethanol gives 38.8 mg. The solid is dried at 20°C and 0.01 mm of Hg for 18 hours and then equilibrated with atmospheric moisture for 24 hours, giving 40.9 mg of the title conpound.

Example 148 [3S-[3a(2),46]]-3-[ [(2-Aaino-4-thiazolyl)f (1carboxy-1 -methyl ethoxy) imino ] acetyl lanino 1 -A-nethvl-210 qxq-1-azetidinesulfonic acid, disodium salt A 3.00 g sample of [3S-[3o(Z),4e]]-3[ [ (2-Amino—5-thiazolyl) - [ (1-caxboxy-l-methyle thoxy) imino] acetyl ] amino] -4-methyl-2-OJ®-lazetidinesulfonic acid, (see example 146), is suspended in 30 ml of water and titrated with IN sodium hydroxide requiring 12.0 ml to give the title disodium salt. The pH is reduced to 6.5 by the addition of a little Dowex 50W-X2 (H+). The mixture is filtered and the filtrate diluted to 66·3 9 with water. A 6.63-g portion is removed for other purposes. The remaining filtrate is lyophilized, giving 2.38 g of solid. Partial equilibration (24 hours) vith atmosphereic moisture gives 2.54 g of the title compound.

Examples 149-151 Following the procedure of—Example-138, but substituting-the compound listed in column I for (Z) -2-amino-o-(methoxyimino)-4-thiazoleacetic 3q acid, yields the compound listed in column II. -17251333 Colum X Opium 11 V Λ X . χ β» Ο I ri rt IL· £17 Π »?ίΐί m Nrt V ri Z I rt rt rt np1 am g X rt ri ml 4 IH iS4lt JL C g «1 | 4r rt 3 rt Λri~ >« a a rt fi 3 « H >, rt rt £?SS& β «π · u fiitix I rt fi rt · ssps m a flirt a a c a rt >* . _ I O ri X rt rt p a rt >, < a >· e e· sIa S A=li -Si & I rt | r» v « » — rt Q v — rt 6 rt -SV I 0 rj rt rt rt Λ3ί2 •m ι · «μ a-3 • Orta rsss £A23 XAA3 78 8 8 «IV I ί^4 to X >»rt P rt C ·»« a o V to rt tt rt 9 « c* a Ai5 V il —^3 ι — X O VS A37 · tt rt JS rt d’l «Χ 7Vv «^is — rt a a U x rt o a ι o V Π tt *— a ό — X rt 72,8 tt rt & X rt rt Ο I a rtm U To a e a tt rt e «I 8 == s 7 SA BOO rue *» X rt £85 IS 8 £1 5 = u 8,5 ?! , 8 ’· AS • M tt C Ai >«—’ 58 T3 tt a rt· 0 I N ? 5 v 51333 -173Example 152 [3S- [3a (2) ,4α] ]-3-I [ (2-Amino-4-thia2Qlyl) [ (1-diphenylmethoxy ear ponyl-1-methyl ethoxy) imino] acetyl] amino] -4-methyl2-oxo-l-azetidinesulfonic acid, potassium salt (1:2) A) [3S-[3α (Z) ,4a] ]-3-[I (2-Araino-4-thiazolyl)[ (l-diphenylmethoxycarbonyl-l-methylethoxy) imino]acetyl ] amino) -4-methyl-2-oxo-l-azetidinesulfonic acid, potassium salt A solution of 440 mg of (Z)-2-amino-o[ (1-carboxy-l-methylethoxy) imino]-4-thiazoleacetic acid and 153 mg of N-hydroxybenzotriazole monohydrate in 3 ml of dimethylformamide is treated with 206 mg of dicyclohexylcarbodiimide. The mixture is stirred at room temperature for 30 minutes under nitrogen and a solution of 180 mg of (3S-cis) -3-amino-4-oethyl-2-oxo-l-azetidinesulfonic acid, (see example 137) and 0.14 ml of triethylamine in 2 ml of dimethylformamide is added (an additional 1 ml of dimethylformamide is used for rinsing) and'the mixture is stirred for about 16 hours. The slurry is evaporated in vacuo and triturated with 12 ml of acetone.

The slurry is filtered and the solid washed with acetone (two 3 ml portions). The combined filtrate and washings are treated with 338 mg of potassium perfluorobutanesulfonate. Dilution with 30 ml of ether gives a gummy solid, which slowly solidifies. The solid is filtered and 3q washed with ether to give 656 mg of the title -174B) [3S-f3a(Z),4a](2-Anino-4-thlazolyl)( (l-carboxy-l-methylethoxy) imino]acetyl]amino]-4methyl-2-oxo-l-azetldineaulfonic acid, potassium «alt (It 2) λ «lurry of 656 mg of {3S-I3a(Z),4a]J-ΙΙ [(2-Aaino-4-thiazolyl)I(1-diphenylmethoxycarbonyl1-aethylathoxy) imino]acetyl]amino]-4-methyl-2oxo-l-azetidinesulfonic acid, potassium salt in 2.3 ml of distilled anisole is cooled to -12°C and 11.5 ml of trifluoroacetic acid (precooled to -10°C) is added. The solution is stirred for 15 minutes and 46 ml of ether followed by 23 ml of hexane are added. After 5 minutes at -10°C and stirring for 15 minutes at rocn temperature, the solid is filtered and washed with ether to give 457 mg of a very hydroscopic gum. This is dissolved in 6 ml of cold water and immediately adjusted to pH 5.6 with 0.4» potassium hydroxide solution. The solution is applied to a 200 ml HP-20 resin and eluted with water. Fractions (50 ml each) 7-11 are combined aad lyophilized to give 239 mg of the title compound as a solid.

Anal. Calc'd. for c13H15OeH5S2K2*1/2 HjO: C, 29.99, H, 3.10? », 13.45, S, 12.32 Found: C, 29.94; H, 3.30; N, 13.30; S, 11.93 NMR(D20) 1.44 (3H d, ;-75), 1.46(6H,S), 4.48(lH,d of t j"75, 5.5), 5.34(1H, d, j»5.5) 6.96ppm(lH,S). 513S3 -175Example 153 (t)-3-Amino-4,4-dimethyl-2-oxo-l-azetidinesulfonic acid A) (-)-4 «^-Dimethyl-Z-oxo-l-azetidine-tertbutyldiphenylsilane A solution of 40.5 ml of t-butyIchlorodiphenylsilane in 112 ml of dimethyl formamide is cooled to 0°C. To this is added 22 ml of triethylamine.

A solution of 12.87 g of 4,4-dimethyl-2-azetidinone in 25 ml of dimethylformamide is added dropwise over 10 minutes to the cooled triethylamine solution. The resulting cloudy solution is stirred for 18 hours at 5°C under argcn. This mixture is poured into 400 ml of ice water and extracted with three 150 ml portions of 2:1 ether:ethyl acetate. The combined extracts are washed with four 100 ml portions of 0.5M monobasic potassium phosphate buffer, one 150 ml portion of sodium bicarbonate solution, two 150 ml portions of water and one 150 ml portion of saturated sodium chloride solution. The solution is dried over sodium sulfate and concentrated in vacuo to yield 33.03 g of the title compound as a solid.

B) (t)-3-azido-4,4-dimethyl-2-oxo-l-azetidinetertbutyldiphenylsilane A solution of 4.25 ml of 1.6M (in hexane) n-butyllithium and 11ml of dry tetrahydrofuran is prepared at -50°C under argon in a 100 ml three-necked flask. A solution of 0.083 g of triphenylmethane in 1 ml of tetrahydrofuran is -176added. The resulting solution is cooled to -60°C, and 1.0 ml of diisopropylamine is added dropwise by syringe. This is stirred for 15 minutes and then cooled to -7S°C. λ solution of 2.3 g of (±)-4,4-dimethyl-2-oxo-l-azetidine-tertbutyldiphenylsilane in 8 ml of tetrahydrofuran is added slowly by syringe. The resulting solution is stirred for 20 minutes at -78°C, during which time heavy precipitation occurs and uniform stirring becomes difficult, λ solution of 1.33 g of ρ-toluenesulfonyl azide in 5 ml of tetrahydrofuran is added dropwise. The resulting mixture is allowed to stir at -78°C for 20 minutes, and 2 ml of trimethylsilyl chloride is added dropwise.

The reaction mixture is warmed to ambient temperature and stirred for 1 hour. Then the mixture is cooled to 0°C and poured into 150 ml of 0°C ethyl acetate. Enough'0.5 M monobasic potassium phosphate buffer is added to make both the aqueous and organic layers clear. The two layers are separated and the organic layer is washed with three 150 ml portions of 0.5 M monobasic potassium phosphate solution, one 150 ml portion of sodium chloride solution, one 150 ml portion of saturated sodium chloride solution and dried over sodium sulfate.

The solution is concentrated in vacuo to 2.83 g of oil, which upon trituration with hexane yields 1.67 g of the title compound as a solid. 177C) (1)-3-Azido-4,4-dimethyl-2-oxo-l-azetidine In a 50 ml three-necked flask, 1.52 g of (i)-3-azido-4,4-dimethyl-2-oxo-l-azetidinetertbutyldiphenylsilane is dissolved in 25 ml of acetonitrile. To this stirred solution is added 0.25 ml of 48« hydrofluoric acid. This is stirred at ambient temperature, and 0.5 ml portions of 48« hydrofluoric acid are added every 60 minutes until, after 6.5 hours, a total of 3.25 ml of 48« hydrofluoric acid has been added.

The reaction mixture is then cooled to 0°C, neutralized with saturated sodium bicarbonate, and extracted with 120 ml of ethyl acetate. The organic layer is then washed with 100 ml of water, 100 ml of saturated sodium chloride solution, and dried over sodium sulfate. The dry solution is concentrated in vacuo to yield 1.34 g of oil.

This impure oil is chromatographed on 27 g of silica gel with hexane, followed by 33« ethyl acetate in hexane, to yield 0.358 g of the title compound as a solid.

D) . (-)-3-Azido-4,4-dimethyl-2-oxo-l-azetidinesulfonic acid, tetrabutylammonium salt To 0.100 g of (±)-3-azido-4,4-dimethy1-2oxo-l-azetidine at 0°C is added under argon 2.8 ml of 0.5M dimethylformamide-sulfur trioxide complex. This mixture is allowed to warm to ambient temperature and stir for 45 minutes. 3Q The solution is then poured into 20 ml of 0.5 M pH 5.5 monobasic potassium phosphate buffer. -170This is washed with three 20 al portions of methylene chloride (discarded) and 0.237 g of tetrabutylammonium hydrogen sulfate is added to the aqueous solution. This was extracted with four 20 al portions of methylene chloride and the combined organic extracts are washed with. ml of 8% sodium chloride solution. The methylene chloride solution is dried Csodium sulfate) and concentrated in vacuo to yield 0.31 g of oil, which appeared by nmr to be 50« dimethylformamide and 50« of the title compound.

E) (—)-3-Amino-4,4-dimethy1-2-oxo-l-azetldinesulfonic acid λ solution of 0.155 g of (±l-3-azido-4,4dimethyl-2-oxo-l-azetidinesulfonic acid, tetrabutylammonium in 0.6 m 1 of methanol is hydrogenated over 10« palladium on charcoal for 20 minutes at 1 atmosphere. The catalyst is filtered off and rinsed with methylene chloride which is combined with the methanol solution. This clear solution is treated with 0.123 ml 97« formic acid. Upon addition of the acid the solution iaeaediately becomes cloudy. After standing for 1 hour at 5°C, the solid.is filtered off to yield 0.0664 g of the title compound, m.p. 200-202*C(dec.).

NMR(D20) 1.64(3H,S), 1.68(3H,S), 4.42(1H,S), IB(KBr) 1765cm1. -179Example 154 [3t(2) ]-3-[[(2-Amlno°4-thiazolyl) (methoxyimino)acetyl]amino]-4 ,4-dimethyl-2-oxo-l-azetidinesulfonic acid, potassiura salt A eolution of N-hydroxybenzotriazole hydrate (50 mg) and (Z)-2-amino-a-(methoxyimino)-4thiazoleacetie acid (0.323 mmole) in 0.5 ml of dimethylformamide is treated with 67 mg of dicyclohexylcarbodiimide under argon at ambient temperature. The resulting mixture is stirred for 1 hour at which time (±)-3-amino-4,4-dimethyl2 -oxo-l-a zetidinesulfonic acid (57 mg; see example 153} is added as a solid followed by triethylamine dropwise (0.05 ml).

The reaction is stirred at ambient temperature for 16 hours. The dimethylformamide is removed under high vacuum at 30°C, and the residue is slurried in 4 ml of acetone and filtered. The filter cake is washed with an additional 4 ml of acetone, and potassium perfluorobutanesulfonate (85 mg) is added to the filtrate, followed by ether. Trituration of the resulting gum with ether gives 40 mg of a tan solid which is chromatographed on a 70 ml HP-20AG column. Elution with water gives 20 mg of the title compound in fractions (5 ml) 16-40 after evaporation, trituration with 1:1 acetone-hexane and drying, m.p. 225°(dec).

Anal. Calc'd. for CjJJ^NgOgS^X: C, 31.80; H, 3.40; N, 16.86; S, 15.43 Found: C, 29.47; H. 3.48: N. 14.98: S, 13.35 -180Example 155 (ί) -4,4-Dimethyl-2-oxo-3- [ (phenylacetyl) amino]1-azetidinesulfonic acid, potassium «alt λ solution of N-hydroxybenzotriazole hydrate (45 mg) and phenylacetic acid (40 mg) in 0.5 ml of dimethylformamide is treated with dicyclohexylcarbodiimide (61 ag) under argon at ambient temperature. The resulting mixture is stirred for 1 hour and (i)-3-aaino-4,4-dimethyl-2-oxo1° 1-azetidinesulfonic acid, (52 ag; see example 153) is added as a solid, followed by triethylamine dropwise (0.04 ml). The reaction is stirred at ambient temperature for 24 hours.

The dimethylformamide is removed under high vacuum at 30°C and the residue is slurried in acetone and filtered. Potassium perfluorobutanesulfonate is added to the filtrate, ether is added and the mixture is cooled. The resulting solid is washed with acetone, hexane, and dried yielding the title compound as a powder.

Anal, calc'd. for Cj^H^jNjOjSK: C, 44.55; H, 4.32; N, 8.00; S, 9.15; K, 11.16 Found: C, 43.83; H, 4.16; N, 7.96; S, 8.76; X, 11.43 NMR(D2q) 1.33(S,3H), 1.58(S,3H), 3.68(S,3Hj, 4.70 (S,1H), 7.56ppm (broads, 5H).

Example 156 (3S-trans)-3-[[(2-Amino-4-thiazolyl)oxoacetyl]amincj-4-methyl-2-oxo-l-azetidinesulfonic acid, potassium salt To a solution of diphenylphosphinyl chloride (1.85 g) in dry dimethylformamide (15 ml) cooled in an ice-methanol bath (-15° to -20°C) is added (2-amino-4-thiazolyl)glyoxylic acid, triethylamine Si 333 -181salt (2.14 g). After stirring for 0.5 hour a solution of (3S°trans)-3-amino-4-methyl-2-oxol-azetidinesulfonic acid (1.08 g; see example 139) and triethylamine (1.92 ml) 3 in dry dimethyl formamide (5 ml) is added to the cold mixed anhydride solution and the reaction mixture is stirred at 5°C for 24 hours. Solvent is removed in vacuo, the residual dark oil is dissolved in wafer, and chromatographed on Dowex 50 £ 2-400 resin (K® form, 200 ml). Upon elution with wafer (15 ml fractions) the crude product is collected in fractions 13-27 (3.37 g). Chromatography on BP-20 resin (200 ml), eluting with water (15 ml fractions), gives the desired product in fractions 18-26. Removal of water in vacuo gives the title compound as an amorphous powder.

Anal. Calc'd for CgHgNjOgS^ (372.42): C, 29.02; H, 2.44; N, 15.04; S, 17.22; K, 10.50 Found: C, 28.87; H, 2.62, N, 14.85; S, 15.09; K, 10.81 Example 157 [3S (R*) 1 -3- [ [ [ (Aminoacetyl) amino] phenyl aeetyl lamino] 2-oxo-l-azetidinesulfonic acid, potassium salt, trifluoroacetate (1:1) salt The deprotection of I3S (R*) J-3-JIIIIII (.4me thoxyphenyl) methoxy] carbonyl ] amino ] acetyl] amino] phenylacetyl] amino]-2-oxo-l-azetidinesulf onic acid potassium salt (see example 127) using trifluoroacetic acid and anisole yields the title compound, melting point 165°C, dee. -182Exawple 158 (3S-trans)-3-Methoxy-4-methyl-2-oxo-3-[[(phenylmethoxy) carbonyl]amino]-1-azetldlneaulfonic acid, potassium salt A) (35-trans)-4-Methy1-3-methoxy-2-oxo-3 [ [ (phenylmethoxy) carbonyl] amino) azetidine A eolation of 2.5 g (0.0106 mole) of (3R-trans) -4-methyl-2-oxo-3 ( [ (phenylmethoxy) carbonyl] amino] azetidine (prepared from d'threonine in 12.6% yield essentially as described for the racemic cis isomer in Example 98C) in 112 ml of 4% borax in methanol is cooled to 0°C and 3.5 ml of t-butyl hypochlorite is added. After 20 minutes the solution is poured into 1 liter of cold water and extracted with two 750 ml portions of cold ethyl acetate. The organic layer is washed with cold water (two 750 ml portions), saturated salt, dried and evaporated to give 3.05 g of crude »,N’-dichloroamide.

A solution of 426 ag of lithium methoxide in 20 ml of dry methanol is cooled to -78°C and diluted with 40 ml of dry tetrahydrofuran. Over 30 seconds a solution of the above chloroamide in 20 ml of tetrahydrofuran (-78°C) are added via syringe. After 20 minutes at -78°C, ml each of acetic acid and trimethyl phosphite are added. After 40 minutes at room temperature the solution is poured into 500 ml of water and extracted with ethyl acetate (two 300 ml portions). The organic layer is washed with -183water, dried, and evaporated to give an oil. Chromatography on a 200 ml silica gel column eluting with 3:1 chloroform-ethyl acetate gives a total of 1.25 g of the title compound.

B) (3S-trans)-3-Methoxy-4-methyl-2-oxo~311 (phenylmethoxy) carbonyl ]amino-l-azetidinesulf onic acid,potassium salt A solution of 800 mg (0.00303 mole) of 10 (3S-trans) -4-methyl-3+aethoxy-2-oxo-4-ί ί (phenylmethoxy) earbonyl] amino]azetidine in 2 ml of dimethylformamide is cooled to 0°C and 4 ml of dimethylformamide-sulfur trioxide complex is added. After 1 hour at 0°C and 4 hours at room temperature the solution is poured into 80 ml of 0.5 M monobasic potassium phosphate (adjusted to pH 5.5) and extracted with methylene chloride (two 50 ml portions, discard). The aqueous layer is treated with 1.04 g of tetrabutyl ammonium sulfate and extracted with dichloromethane to give 1.42 g of oil. This is dissolved in acetone and treated with 1.04 g of potassium perfluorobutanesulfonate in 10 ml of acetone. Dilution with 250 ml of ether and extensive trituration of the oily solid gives 584 mg of crude product. Chromatography on BP-20 AG (200 ml) gives 418 mg of purified product in fractions (100 ml) 13-16 (elution with 1 liter of water and then 9:1 water-acetone). Trituration of 114 mg of this material with ether gives 104 mg of an analytical sample.

Analysis calc'd for C^gH^^NjO^SK-HgOs C, 39.06? H, 4.04; N, 7.01; S, 8.03; K, 9.78 Pound: C, 38.91; H, 3.62; N, 6.91; S, 8.06; X, 9.51 35 NMR(D2Q) 1.33(38, d, ?«7), 3.46(3H,S), 4.22(2H, d of d, ;"6), 5.18 (2H,S), 7.43ppm(5H,S). -184gxample 159 (35-trans)-3-Methoxy-4-methyl-2-oxo-3-((phenylacetyl)amino]-l-azetldinesulfonic acid, potassium salt (3S-tran>) -3-amino-3-MethDxy-4-sethyl-2-O3®_l_ azetidinesulfonic acid, tetrabutylanmonium salt is prepared by the catalytic hydrogenation of (3S-trans)-3-methoxy-4-methyl-2-oxo-3-1[(phenylmethoxy) carbonyl]amino]-1-azetidinesulfonic acid, potassium salt (see example 158) after conversion to the tetrabutylannonium salt. Following the procedure of example 88, bit utilizing (3S-tons)-3-anino-3-methQxy4-methyl-2-oxo-l-azetidinesulfonic acid, tetrabutylammonium salt and phenylacetyl chloride yields the title compound.

Anal, calc'd for C13H1SN2O6SX: C,42.61; H,4.31j N,7.65 Found: C,39.67; H,4.09; 11,7.30 IWR (D20) 1.29(BH, d, j-7) 3.45(3H,S), 3.73(2H,S), 4.36 (2H, d of d, j-6), 7.38ppm (5H,S).

Example 160 [3S-[3a(Z),46]]-3-[I(2-Amino-4-thiazolyl)[[2(diphenylmethoxy)-2-oxoethoxy]imino]acetyl]amino]-2-aethyl-4-oxo-l-azetidinesulfonic acid, potassium salt Following the procedure of example 138, but substituting (Z)-2-amino-a-[[2-(diphenylmethoxy)2-oxoethoxy]imino]-4-thiazoleacetic acid for (Z)-2-amino-a- (methoxyimino) -4 -thiazoleacetic acid and first treating the (3S-cis)-3-amino4-methyl-2-oxo-l-azetidinesulfonic acid, with triethylamine, yields the title compound, melting point 155-160°C, dee. 185Examale 161 [3S-[3o(Z),4S]]-3-I[I(Carboxymethoxy)imino](2-amino4-thiazolyl) acetyl]amino]-4-methyl-2-oxo-lazetidinesulfonic acid, dipotassium salt 5 The deprotection of I3S-I3o(Zl,4aJJ-3-JJt2euaino-4-thia zolyl) I [2-(diphenylmethoxy)-2-oxoethoxy]imino] acetyl ] amino] -2-methyl-4 -oxo-l-a zetidineulfonic acid, potassium salt (see example 160) using trifluoroacetic acid and anisole yields the title compound, which decomposes at:>250°C.

Example 162 (S)-3-I[I(2,6-Dichloro-4-pyridlnyl)thio]acetyl]amino]-2-oxo-l-azetidinesulfonic acid, potassium salt Acylation of (S)-3-amino-2-oxo-l-azetidinesulf onic acid, tetrabutylammonium salt (see example 6A) with [(2,6-dichloro-4-pyridinyl)thio]acetic acid, 4-nitrophenyl ester, followed by treatment with potassium perfluorobutane sulfonate, yields 2o the title compound, melting point 212-214°C.

Example 163 I3S(R*)] -3-1 HI (4-Amino-2,3-dioxo -1-piperaz inyl) carbonyl]amino]phenylacetyl]amino]-4-methyl-225 oxo-1-azetidinesulfonic acid, potassium salt [3S(R*)]-]4-II[2-1(4-methyl-2-oxo-l-sulfo3-azetidinyl) amino]-2-oxo-l-phenylethyl] amino]carbonyl]-2,3-dioxo-l-piperazinyl]caxbamic acid, phenylmethyl ester, potassium salt (see example 149) is hydrogenated using gaseous hydrogen and 10% palladium on charcoal as catalyst, yielding the title compound, melting point 165°C, dec. 5x323 -186Example 16« [38(Ζ))-3-[I(2-Amino-4-thiazolyl)][(1-carboxy-lmethylethoxy) iaino]acetyllamino]-2-oxo-l-azetldinesulfonic acid, sodium salt (1:2) [3S(Z)]-3-I[(2-Amino-4-thiszolyl)[]2-(diphenylnethoxy)-1,1-dimethyl-2-oxoethoxy]iaino]acetyl)amino]2-oxo-l-azetidinesulfonic acid, tetrabutylammonium salt (see example 28) is deprotected using trifluoroacetic acid and anisole to yield the title compound, melting point 185°C, dee, after conversion to the disodiua salt with aqueous sodium hydroxide and purification on BP-20.

Examples 165-168 Following the procedure of Example 11, but substituting the acid listed in column X for (I ] -2-amino-a-1 [hydroxy (phenylmethoxy) phosphinyl ] methoxy]imino]-4-thiazoleacetic acid, yields the compound listed in column XI. -187I »4 0 ·* >1 X Al 1 e ι 1 0 to ϋ in 0 © PM £ 1 fl rt PM x e ο» ι 9 CM (Q ι β e to Wrt fi 1 — rt 1 <» — 0 fl Ό *4 t to o ® e — 0 tu A Oh AJ rt 1 fi Q · 0 3 —’ fi rt fi —b A) C-4 1 c-Η fl o fi rt a —4 >1 © © HH gAJ 0 ή a rt g OT 0 X N g >O fl 0 *0 E o >4 © Gt O fl e-H— o. A fl H-C X 1 ·* H Ert » -* Al 0 >-H O' A to A O-H > *U rt 0 Ν ΧΌ C © 1 rt U4 ki Α Ό0 > a 0 -η -4 E —· fl 1 >*©*H© H 0 rt A) A) A> won » 0,0 ON H N * · a a ©rt >e _ — — fl fl PM CO © A>X N © cw g 1 Ort 1 g © rt © 1 0 fl g © E 9 . οι ι >uw a χ ο ό X X fl Ή i cn c-h to g >4 fl 1 0 to *» ο.—»n > -© CN rt X 0 1 j A ~to ® XdX o 0 0 ou fi tt· 0 c-4 —* > fl · 0 M 0* u A -HO rt ί X fl "OA 0 W 0— rt fi Al C © g — o a — NO© Ό »0 0 9 to © 0 © a > 1 C QiO >-h eno g tu rt 1 XN s PM © x u-h © a rt rt pm 0 3 — X » * ι >E e Ό ? A> — X—rt —— m Gc fl rt O' 1 Q fi — Al 0 e 1 W Ή0 ——Ό e © ©rt *"* ©SO · m >, u m — rt rt rt rt » C 0 1 grt fl o 1 J= β · — >>Al Al PM *4 CU PM rt g AS © 0 *3 v — fi fi ©rt *-0 X ffl Ο n — rt ON© — rt 01 —. fi— Sc 0 E-^w 1 NX fl g f A> fi —> © rt * 1 — c rt fl Μ I (*) ©rt NX > *U WOO*1 I H flrt - I N A ** QtAl Ό0 ι e — © 0 1 A «—- fl rt WH 0-40 λΜ rt rt CO a— 0 rt w ι © to Ό O © m ta E 9 o rtrt rt X C — rt £ ·«* fl fl rt **9(8 0« — Oc >i 0 O I 4> ί 1 B 1 0 to fi rt — rt1 Ί >iO >1 rt 1 g •0 fi fi CO fl rt ©rt Ort Β Ό — 0 X £ X O C 1 rt fl Girt W fl on >» -H P"R fl tw * C 0 Ό >1 ϋ □ 1 (M Q rt rt *- ΧΌ — rt 0 1 Ort >0 — o wo e N AS O X N 1 Ό Art e — 9 fl o e PMrt 0 O § © ‘"’X x © 1 Wrr « rt X 1 0 o XX 0 >» Ο X rt © — X Gt>iO CJ X 1 0 AS grt 0 — 1 rt 0 0 © rt W rt to AS X C >X Ό > 1 © Al rt a e a >CCU Φ ε ** © © V X rt rt fl g 3 >,rt χ ε 1 Ν Ό © rt 1 X 0 Gt— in fl-π c Ό PM ο N — 0 — w £ © I 1 X fl — c — ©rt N to «—< A> rt —rt — G. U C Ν © X8 € 1 rt >,© PM — g AS PM fl β CU CUXee . in to P* CD to to to to rt rt rt rt -188Example 169 (trans)-3-Araino-4-ethyl-2-oxo-l-a2etidinesulfonic acid A) t-Boe-N-methoxy-B-threoethylserinamide threo-D,I,-S-Ethylserine (1.33 g) is dissolved in 10 nl of 2N potassium hydroxide and 5 al of t-butanol. After the addition of 2.46 g of di-t-butylpyrocarbonate the two-phase mixture is stirred for 4 hours at aafeient temperature. O-Methylhydroxylammonium chloride (1.25 g) is added and the pH is adjusted to 4 with IN hydrochloric acid. l-Ethyl-3-(3-dimethylaminopropyl ) carbodi imide, hydrochloride (1.92 g) is added and the pH is again adjusted to 4. After stirring for 1 hour the reaction mixture is saturated with sodium chloride and extracted with four 50 ml portions of ethyl acetate.

The ethyl acetate extracts are combined and dried over MgSO*. Removal of the solvent in vacuo yields 1 g of the title compound.

B) t-Boc-O-methanesulf onyl-N-methoxy- β-threopropionamide t-Boc-N-methoxy-S-threoethylserinaaide (10.5 g, is dissolved in 65 ml of pyridine. Methanesulfonyl chloride (4.65 ml) is added dropwise at 0°C.

After stirring for 3 hours at ambient temperature, the reaction mixture is poured into 200 g of ice -and 300 ml of IN hydrochloric acid. The pH is adjusted to 4 with concentrated hydrochloric aeid. After extraction with three 85 ml portions 51333 -189of ethyl acetate the combined extracts axe dried over MgSO^ and concentrated in vacuo. The residue is treated with carbon tetrachloride and concentrated again. Stirring with ether followed by filtration yields 6.9 g of the title compound.

C) (trans) -3-t-Butoxycarbonylamino-4-ethyl-lmethoxy-2-azetidinone Anhydrous potassium carbonate (4.15 g) and 125 ml of dry acetone are brought to reflux and 3.4 g ef t-Boc-O-methanesulfonyl-N-methoxyB-threo-propionamide in 25 ml of acetone is added. After 1 hour the reaction mixture is cooled and filtered, and the filtrate is concentrated in vacuo. The oily residue is stirred with hexane to yield 2.2 g of the title compound.

D) (trans)-3-t-Butoxycarbonylamino-4-ethyl-2azetidinone (trans)-3-t-Butoxycarbonylamino-4-ethyl-lmethoxy-2-azetidinone (3 g) is added to 170 ml of liquid ammonia at -78°C under nitrogen and 1.68 g of eodium is added in 5 portions with stirring over a 5 minute period. Stirring is continued for 30 minutes. ' Ammonium chloride is then added slowly until the blue color of the reaction mixture disappears. After removal of the ammonia under nitrogen the solid is extracted with two 100 ml portions of ethyl acetate.

Removal of the solvent followed by drying in vacuo yields 2.7 g of the title compound. 51333 -190E) (trans)-3-t-Butoxycarbonylamino-4-»thyl-2oxo-l-azetldinesulfonlc acid, tetrabutylammonium salt To a 2 ml of absolute pyridine in 20 ml of dry dichloromethane is added trimethylsilylsulfonyl chloride (3.7 ml) in 5 ml of dry dichloromethane. The addition is accomplished at -30°C, under nitrogen, over a 10 minute period. After stirring at ambient temperature for 30 minutes, the flask is evacuated to yield a pyridine-sulfur trioxide complex, (trans)-3t-Butoxyeazbonylamino-4-ethyl-2-azetidinone (2.67 g) and 20 ml of dry pyridine are added to the flask which is then placed in an oil bath preheated to 90°C. After 15 minutes a clear solution is obtained and poured into 200 ml of a IB solution of dibasic potassium phosphate. After the addition of 27 g of dibasic potassium phosphate and 100 ml of water a clear solution is obtained. The solution is extracted with two 60 ml portions of ethyl acetate. Tetrabutylammonium hydrogensulfate is added to the agueous layer, and the agueous solution is extracted with three 100 ml portions of dichloromethane and the combined organic layers are dried over MgSOj. Concentration in vacuo yields 6.9 g of the title compound. -191F) (trans) -3-Amino-4-ethyl-2-oxo-l-azetidinesulf onic acid (trans)™3-t-Butoxycarbonylamino-4-ethyl-2oxo-l-azetidinesulfonic acid, tetrabutylammonium salt (6.75 g) in 40 ml of 98% formic acid is stirred for 3 hours at ambient- temperature. Dichloromethane (60 ml) is added and the mixture is refrigerated for about 16 hours. The resulting precipitate is separated by filtration and then 1® dried in vacuo to yield 0.85 g of the title compound, melting point 185°C, dec.

Example 170 (trans,2)-3-11(2-Amino-4-thiazolyl)[(1-carboxy15 1-methylethoxy)imino]acetyl]amino]-4-ethyl-2oxo-1-azetidinesulfonic acid, dipotassium salt A) (trans,Ζ)-3-Π (2-Amino-4-thiazolyl)[(1diphenylmethoxycarbonyl-l-methylethoxy) imino]20 acetyl]amino]-4-ethyl-2-oxo-l-azetidinesulfonic acid, dipotassium salt (trans)-3-Amino-4-ethyl-2-oxo-l-azetidinesulfonic acid (0.55 gl and 335 mg of triethylamine are dissolved in 50 ml of dry dimethylformamide. (2)-2-Amino-e-[(1-diphenylmethoxycarbonyl-1methylethoxy)imino]-4-thiazoleacetic acid (1.14 g) is added with stirring at 0°C followed by 450 mg of hydroxybenzotriazole and then 0.69 g of dicyclohexylcarbodiimide. After stirring for 30 about 16 hours at 0°C the flask is evacuated.

Dry acetone (25 ml) is added to the solid with 51333 -192stirring. The mixture ie filtered and 0.94 g of potassium perfluorobutanesulfonate is added to the filtrate followed by 100 ml of ether.

After standing for 1 hour at 0°C the solid is filtered, washed with ether and dried in vacuo yielding 1.58 g of the title compound.

B) (trans, Ζ) -3-11 (2-Amino-4-thiazolyl) [ (1-carboxy1-methylethoxy) imino] acetyl] amino] -4-ethyl-2oxo-l-azetidlnesulfonie acid, dipotassium salt To a suspension of 1.31 g of (trans],Z)-3II(2-amino-4-thiazolyl)] (1-diphenylmethoxycarbonyl1-methylethoxy! imino] acetyl] amino] -4- After stirring for 2 hours at -10°C a clear solution is obtained. At -30°C 80 ml of dry ether is added and the resulting precipitate is filtered and then treated with 5 ml of water.

The pH is adjusted to 5.5 with IN potassium hydroxide at 0°c, and the mixture is filtered to remove unconverted starting material. The filtrate is chromatographed on HP-20 with water as eluent. Lyophilization yields 185 rag of the title compound, melting point 160°C, dec. 513 3 3 -193Examplc 171 i3S(Z)]-3°]f (2-7ttalno-4-thiazolyl) [(4-hydroxy-4oxobutoxy) imino] acetyl 3 amino] -2-oxo-l-azetidinesulfonic acid, potassium salt § The deprofceetion of ]3S(Z)]-3-II(2-amino-4thiazolyl) [J4= (diphenylmethoxy) -4-oxobutoxy]imino]acetyl] amino] -2-oxo-l-azetidinesulfonic acid, potassium salt (Bee example 166) using trifluoroacetic acid and anisole yields the title compound, melting point >200°C.

Example 172 (S)-3-J (2-(Aminomethyl? benzoyl] amino]-2-oxo-lazetidinesulfonic aeid, inner salt The deprotection of (S)-2-oxo-3-[Γ2-ΙΠ(phenylme thoxy) carbonyl] amino]methyl]benzoyl) amino] -1azetidinesulfonic acid, potassium salt using hydrogen gas, palladium on charcoal, and hydrochloric acid, yields the title compound, melting point 162-165°C.

Example 173 (S)-3-[I [2-(4-Fonayl** 1-piperazinyl)-5-hydroxypyrido [2,3-d]pyrimidin-6-yl]carbonyl lamino]-225 oxo-l-azetidinesulfonic acid, potassium salt (S)-3-Amino-2-oxo-l-azetidinesulfonic acid, tetrabutylammonium salt (see example 6A) is coupled with 2-(4-f ormyl-l-piperazinyl)-5-hydroxy6-1 (4-nitrophenoxy)carbonyl]pyrido (2,3-d]pyrimidine and treated with potassium perfluorobutanesulfonate in acetone to yield the title compound, melting point 290°C, dec. -194Example 174 (3S-trans)-a-(I(4-Mcthyl-2-oxo-l-sulfo-3-azetidinyl)amino]carbonyl]benzeneacetic acid, dipotassium salt (3S-trans)-3-Amino-4-aethyl-2-oxo-l-azetidinesulfonic acid (see example 139) is coupled with a-(carboxyl)benzeneacetyl chloride and treated with triethylamine and potassium perfluorobutanesulf onate to yield the title compound, melting point 147°C, dec.

Example 175 l3S-toanej-3-Ai*iao-4-cyclohexyl"3"Oxo-l-azetldinesulfonic acid A) a-(t-Butoxycarbonylamino)-8-cyclohexyl-Bhydroxy-threo-propionic acid B-Cyclohexyl-a-amino-B-hydroxy-threo-propionic acid (15 g) is suspended in 150 ml of acetonitrile and 70 ml of water. Triethylamine (17.8 g) is added and the mixture is heated with stirring to 60°C. At this temperature a clear solution is obtained and 21.0 g of di-t-butylpyrooarbonate is added and stirring at 60°C is continued for 1.5 hours. She solvent is removed in vacuo and 50 ml of water is added. The aqueous layer is extracted with ethyl acetate at a pH of 2, which is adjusted by addition of 3N HCl. The organic layer is separated, dried over NajSO^ and evaporated to dryness. The remaining crystalline material is filtered with petrol » ether, yielding 20.4 g of the title compound, melting point 113-115°C. -195B) a-(t-Butoxycarbonylamino)-a-cyclohexyl-ghydroxy-N-me thoxy-threo-propionamide β-(t-Butoxycarbonylamino)-B-cyclohexyl-ghydroxy-threo-propionic acid (20.2 g) and 7.6 g of O-methylhydroxylamine hydrochloride are suspended in 350 ml of water and 175 ml of t-butanol. The pH of the mixture is adjusted with potassium carbonate to 4.l-Ethyl-3-(3-dimethylaminopropyl)carbodiimide CIS.4 g) is added and the pH is maintained at 4 with stirring for 1.5 hours. t-Butanol is removed in vacuo and the remaining agueous solution is saturated with sodium chloride and extracted twice with 100 ml portions of ethyl acetate. The organic layers are combined, dried with Na^SO^ and evaporated to dryness. The remaining crystals are filtered off with petrol ether yielding 18.6 g of the title compound, melting point 125-127°C.

C) a-(t-Butoxycarbonylamino)-B-cyclohexyl-B(methanesulfonyloxy)-N-methoxy-threo-propionamide a-(t-Butoxycarbonylamino)-β-cyclohexyl-Bhydroxy-N-methoxy-threo-propionamide (18.3 g) is dissolved with stirring in 100 ml of dry pyridine. The solution is cooled with stirring to 0°C and 9.3 g of methanesulfonyl chloride is dropped in. After one hour at 0°C an additional 3.3 g of methanesulfonyl chloride is added and stirring is continued for one more hour. The solution is poured into 300 ml of ice water, -196200 nl of ethyl acetate is added and the pH ia adjusted to 3 with dilute sulfuric acid. The organic layer ia separated, dried with Na^O^ and the solvent is removed in vacuo. The remaining solid ia collected with petrol ether yielding 19.0 g of the title compound, melting point 150-152°C.

D) [3S-trans] -3- (t-Butoxycarbonylamino) -4cyclohexyl-l-methoxy-2-azetidinone a-(t-Butoxycarbonylamino)-B-cyelohexyl-8(aethaneaulfonyloxy)-N-methoxy-threo-propionamide (18.7 g) is dissolved in 500 ml of dry acetone. Potassium carbonate (9.8 g) is added and the suspension is heated to reflux temperature with stirring for 5 hours. The insoluble inorganic material is filtered off and the solvent removed in vacuo and the remaining oil is dissolved in 30 ml of ethyl acetate, ϋροη the addition of petrol ether, the title compound precipitates and is filtered off (12.9 g), melting point 110-112°C. Ε) 13S-trans]-3-(t-Butoxycarbonylamino)-4cyclohexyl-2-azetidinone (3S-trans3-3-(t-Butoxycarbonylamino)-4eyelohexyl-l-methoxy-2-azetidinone (1 g) is added to 50 ml of liquid ammonia with stirring. Sodium (0.154 g) ia added in 5 to 6 portions within 5 minutes. After this time an additional -197amount of 0.025 g 0odium is added and stirring is continued for 5 minutes. Ammonium chloride (0.89 g) is added and the ammonia is removed.

The residue is extracted with warm ethyl acetate The organic extract is evaporated to dryness and the remaining crystals of the title compound are filtered with petrol ether, yielding 0.5 g, melting point 130-132°C.

F) 13S-trans] -3- (jj-Butoxycarbonylamino) -4eyclohexyl-2-oxo-l-azetidinesulfonic acid, pyridine salt 135-trans]-3-(t-Butoxycarbonylamino)-4cyclohexyl-2-azetidinone (5.3) is dissolved in ml of methylene chloride and 80 ml of dimethylformamide. After the addition of 60 mmole of pyridine-sulfur trioxide complex the solution is stirred for t hours at room temperature. Removal of the solvent in vacuo 2o yield 11.3 g of the title compound as an oil.

G) 135-trans] -3-- (t-Butoxycarbony 1 amino) -4 cyelohexyl-2-oxo-l-azetidinesulfonic acid, tetrabutylammonium salt I35-trans]-3-(t-Butoxycarbonylamino)-4cyclohexyl-2-oxo-l-azetidinesulfonic acid, pyridine salt (11.3 g) is dissolved in 250 ml of water. Tetrabutylammonium hydrogensulfate (9.0 g) is added with stirring and the pH is 3q adjusted fo 6.5 with IN potassium hydroxide.

The agueous solution is extracted twice with 51333 -198200 «1 portion* of methylene chloride. The organic portion* are dried with Na^SO^, filtered and the solvent ie distilled off, yielding 8 g of the title compound, melting point 135-138°C.

H) I3S-tran«] -3->mino-4 -cycloh exyl-2-oxo-lazetldinerulfonic acid [3S-trane]-3- (t-Butoxycarfaonylamino1-4cyclohexyl-2-oxo-l-azetidinesulfonic acid, tetrabutylammonium salt (3.8 g) ie stirred in 20 ml of formic acid for 3 hours, followed by the addition of 20 ml of methylene chloride.

The precipitated title compound (1.0 g) ie filtered off, melting point 217-219°C.

Example 176 [3S-[3α(Z),4b])-3-(((2-Amino-4-thiazolyl)(methoxyimino)acetyl]amino]-4-eyclohexyl-2oxo-l-azetidinesulfonic acid, potassium salt {3S-transJ-3-Amino-4-cyclohexyl-2-oxo1-azetidinesulfonic acid (0.25 g) is dissolved in 30 ml of dry dimethylformamide and 0.12 g of triethylamine with stirring. When a clear solution is obtained, (Z)-2-amino-a-(raethoxy25 imino)-4-thiazoleacetic acid (0.2 g), 0.16 g of hydroxybenzotriazole and 0.42 g of dicyclohexylcarbodiimide are added. Stirring is continued for 48 hours at ambient temperature. The precipitated urea is filtered off and the solvent is removed in vacuo. The residue is 51333 -199dissolved in 10 ml of acetone and 0.41 g of potassium perfluorobutanesulfonate is added.

After the addition of 50 ml of ether, the title compound precipitates and is filtered.

Column chromatography using HP-20 and water/ acetone (9:1) as eluent, yields 0.36 g of product, melting point 200-205°C (after lyophilization).

Example 177 [3S-[3a(2),4 6]]-3- [ I(2-amino-4-thiazolyl)I(1earboxy-l-methylethoxy) imino] acetyl] amino] -4cyclohexyl-2-oxo-l-azetidinesulfonic acid, dipotassium salt A) [3S-[3a(Z),4B]]-3-[[(2-Amino-4-thiazolyl)[(1diphenylmethoxycarbonyl-l-methylethoxy) imino] acetyl]amino]-4-cyclohexyl-2-oxo-l-azetidinesulfonic acid, potassium salt [35-trans]-3-Amino-4-cyclohexyl-2-oxo-l20 azetidinesulfonic acid (0.2 g; see example 175} is dissolved in 30 ml of dimethylformamide and Q.09 g of triethylamine with stirring. Hydroxybenzotriazole (0.12 g), 0.30 g of (Z)-2-amino-o[ (1-diphenylmethoxycarbonyl-l-methylethoxy) imino] 25 4-thiazoleacetic acid and 0.33 g of dicyclohexylcarbodiimide are added and stirring at ambient temperature is continued for 12 hours. The precipitated urea is filtered off and the mother liquor is evaporated to dryness. The remaining oil is dissolved in 5 ml acetone, treated with 0.3 g of potassium perfluorobutanesulfonate and -200poured into 100 nl of ether with stirring. [3S-[3a (2),46]] -3-11 C2-Xmino-4-thiazolyl) I (1-diphenylmethoxycarbonyl-l -methylethoxy) imino] acetyl] amino] -4-cyclohexyl-2-oxo-l-azetidineaulfonie acid, potassium salt (0.61 g) precipitates and is filtered off.

B) [3S-[3a(Ζ),4 a]]~3~I I(2-Amino-4-thiazolyl)[(1earboxy-l-aethylethoxy) imino] acetyl] amino] -4cyelohexyl-2-oxo-l-azetidinesulfonic acid, dipotassium aalt [3S-[3a(Ζ),4 B]]-3-[I(2-Amino-4-thiazolyl)[ (1 -dipheny lmethoxycarbonyl-l-methylethoxy) imino]aeetyl]amino]-4-cyclohexyl-2-oxo-l-azetidinesulfonie acid, potassium salt (0.61 g) is suspended in 6 ml of anisole, cooled to -15°C and 5 ml of trifluoroacetic acid is dropped in with stirring.

The temperature is maintained for one hour and then lowered to -30°C. About 100 ml of dry ether is added at such a rate, that the temperature does not exceed -10°C. The precipitated compound is filtered off and chromatographed using HP-20 resin and water/acetone (9:1) as eluent, yielding 0.3 g of the title compound, melting point ll-120°C, dec, (after lyophilization). -201Example 178 135-13α, 4 B]-4-Cyclohexyl-3-I[II13-1(2-furanylmethylene)amino] -2-oxo-l-imidazolidinyl] carbonyl]aaino]phenylacetyl]amino]-2-oxo-l-azetidinesulfonic acid, potassium salt [3S-trans]-3-Amino-4-cyclohexyl-2-oxo-lazetidinesulfonic acid (0.1 g; see example 175] is dissolved in a mixture of 30 ml of dry dimethylfozmamide and 0.05 g of triethylamine with stirring. I[II(2-Furanylmethylene)amino]2-oxo-l-imidazolidinyl]carbonyl]amino]phenylacetic acid (0.14 g), 0.06 g hydroxybenzotriazole and 0.17 g of dicyclohexylcarbodiimide are added and the solution is stirred for 5 days at room temperature. The solvent is removed in vacuo, the residue is dissolved in 10 ml of acetone and the precipitated urea is filtered off.

The mother liquor is agitated with 0.15 g of potassium perfluorobutanesulfonate and diluted with 50 ml of ether. The precipitate is filtered, and chromatographed with HP-20 resin using water/acetone (9:1) as eluent yielding 0.14 g of product, melting point 195-200°C, dec, (after lyophiliaztion). -202 Example 179 l3S-l3a (R*r, 13S-trans]-3-Amino-4-cyclohexyl-2-oxo-lazetidinesulfonic acid (0.1 gt sea example 175} ie dissolved in 30 ml of dimethylformamide and 0.5 g of triethylamine with stirring. (R)-o[ ((4-ethyl-2,3-dioxo-l-piperazinyllcarbonyl]aminojbenzeneacetic acid, 0.06 g of hydroxybenzotriazole and a. 17 g of dicyelohexylearbodiimide are added, and tha mixture is stirred for about 16 houre at room temperature, ihe solvent is distilled off in vacuo and the remaining oil is dissolved in 10 ml of acetone. The precipitated urea is filtered off and the mother liquor is agitated with 0.15 g of potassium perfluorobutanesulfonate and diluted with 50 ml of ether.

The precipitate is filtered off and chronatographed with HP-20 resin, using water/acetone (9:1) as eluent, yielding 0.15 g of the title compound, melting point 175*180°C (after lyophilization).

Example 180 (trans,Z)-3- f f ,(3^Amino-4-thiazolyl) (methoxyimino)acetyl)amino2-4-ethyl-2-oxo-l-azetidinesulfonic acid, potassium salt Following the procedure of example 170, part A, but substituting (Z)-2-amino-a-(methoxyimino)-4-thiazoleacetic acid for (Z)-2-amino-aI (l-diphenyboethoxycarbonyl-l-methylethaxy) imino] 4-thiazoleacetic acid, yields the title compound, melting point 190°C, dec. 203Example 181 (t) - (trans) -3-Amino-2-oxo-4-pheny1-1-azetidinesulfonic acid A) (±)-(trans)-2-oxo-4-phenyl-l-azetidine-tertbutyldiphenylsilane A solution of tert-hutylchlorodiphenylsilane (20.56 g] in dimethylformamide (45 ml) is cooled to 0°C under argon and treated vith triethylamine 1° (10.4 ml) and then (£l-2-oxo-4-phenyl-l-azetidine.

After several hours at 0°C, the resulting mixture is treated with additional triethylamine (1 mil and tert-hutylchlorodiphenylsilane (2.11 g), and allowed to stir for 65 hours at 5°C. The reaction mixture is poured into ice water (300 ml) and extracted with 3:1 ether-ethyl acetate (three 125 ml portions). The organic extracts are washed with pH 4.5 phosphate buffer (three 50 ml portions), saturated sodium bicarbonate solution (50 ml), water (two 50 ml portions) saturated sodium chloride solution and dried (Na^SO^). Filtration, and concentration in vacuo yields a solid which is washed with hexane to give after drying (high vacuum) g of the title compound as a solid. -204Β) (ί)-(trans)-3-azido-2-oxo-4-phenyl-l-azetidinetert-butyldiphenylsilane λ 50 nl flask equipped with stirring bar, gas inlet, and septum is flane dried under argon and charged with n-butyl lithium (0.65 ml of a 1.6 M solution in hexane} which is cooled to -40°C and dissolved in tetrahydrofuran (2 ml). Diisopropylamine 10.16 ml) is added dropwise, the resulting mixture is stirred for 30 minutes and cooled to -78°C. A solution of (ii-(trans)2-oxo-4-pbenyl-l-azetidine-tert-butyldiphenylsilane (400 mg) in tetrahydrofuran (1.5 ml) is added dropwise over about 5 minutes. After stirring an additional 20 minutes the solution is treated with p-toluenesulfonyl azide (204 mg) in tetrahydrofuran (0.5 ml). The resulting mixture is stirred 10 minutes at -78°C and treated dropwise with chlorotrimethylsilane (0.4 ml). After an additional 10 minutes of stirring, the cooling bath is removed and the reaction mixture is stirred at ambient temperature for 2.5 hours.

Then, while cooling at 0°C, ethyl acetate (20 ml) is added followed by pH 4.5 phosphate buffer (8 ml). The organic layer is washed with additional buffer (two 8 ml portions), 5« sodium bicarbonate solution (three 10 ml portions), 50% sodium chloride solution (10 ml), saturated sodium chloride solution (10 ml) and dried (Na^SO^). Filtration and concentration in vacuo yields 500 mg of oil which is flash, chromatographed with 5% ethyl acetate-hexane, yielding the title compound (253 mg). 513 93 -20510 C) li)-(trans)-3-azido-2-oxo-4-phenyl—l-azetidine A solution of 17 g of crude Ci)-(trans)-3azido-2-oxo-4-phenyl-l-azetidine-tert butyldipbenylsilane is dissolved in methanol (240 ml) and treated dropwise at 0°C with concentrated HCl (35 ml). The cooling bath is removed, the reaction stirred at ambient temperature for 1 hour, and recooled to 0°C whereupon saturated sodium bicarbonate solution is added to neutrality The resulting mixture is extracted with ethyl acetate (one 300 ml portion and four 100 ml portions) and the organic extracts are washed with 1:1 5% sodium bicarbonate 50% sodium chloride solution, saturated sodium chloride solution, and dried (Na^SO^). Filtration and concentration in vacuo yields 15 g of a heavy oil which is chromatographed on 100 g of silica gel with 20% ethyl acetate-hexane, yielding 460 mg of the title compound.

D) (i)- (trans)-3-azidO"2-oxo-4.-phenyl-1-azetidineT sulfonic acid, tetrabutylammonium salt A solution of (±)-(trans)-3-azido-2-oxo4-phenyl-1-azetidine (300 mg) in dimethylformamide (3 ml) is cooled to 0°C under argon and treated dropwise with a complex of dimethylformamide and sulfur trioxide (4.78 ml of a 0.5 M solution in dimethylformamide). The cooling bath is removed, the reaction mixture is stirred at ambient temperature for 2 hours 51333 -206and poured Into 80 ml of 0.5 M monobasic potassium phosphate (pH 5.5). The solution is extracted with dichloromethane (discarded) and 541 mg of tetrabutylammonium bisulfate ie added. The resulting mixture is extracted with, dichloromethane and the organic extracts are washed with 10« sodium chloride solution and dried OtajSO^l. Filtration and concentration in vacuo affords 8Q0 mg of oil: approximately 40« the desired product, the remainder dimethy lf ormamide. This mixture is used without purification in the next step.

E) (+)- (trans) -3-amino- 2-oxo-4.-phenyl-1 -azetidinesulfonie acid A solution of (ί)-(trans)-3-azido-2-oxo-4—phenyl1-azetidinesulfonic acid, tetrabutylammonium salt in 4 ml of methanol is hydrogenated over 30 mg of platinum oxide at 1 atmosphere and room temperature. After 15 minutes, the system is evacuated and fresh hydrogen is introduced.

After an additional 45 minutes the reaction is complete, and the system is flashed with nitrogen. After several days at room temperature in dichloromethane-methanol (4:1, 200 ml) catalyst aggregation is complete and filtration is accomplished. The filtrate is concentrated in vacuo to 18 ml and 0.2 ml of 97« formic acid is added. After cooling at 5°C for several hours the resulting, solid is filtered and washed with dichloromethane to afford, after drying, 150 mg of the title compound as a solid. Analysis calc'd for C^H^^O^S: C, 44.62; H, 4.17; N, 11.57; S, 13.23 Found: C, 43.36; B, 4.31; N, 11.09; S, 13.02 -207Exarople 182 (±) - (trans)-2-Qxo-4-phenyl-3—I(phenylacetyl)amino]1-azetidinesulfonie acid, potassium salt A) (±)-trans-2-oxo-4-phenyl-3-I(phenylacetyl)amino]1-azetidinesulfonic acid tetrabutylammonium salt A solution of N-hydroxybenzotriazole monohydrate (52 mg) and phenylacetic acid (46 mg) in dimethylformamide (0.3 ml) is treated with solid dicyclohexylcarbodiimide (70 mg) under argon at 0°C. The cooling bath is removed and the resulting mixture stirred at ambient temperature for 1 hour. After dilution with additional dimethylformamide (0.3 ml), (±)-(trans)-3-amino-2-oxo-415 phenyl-1-azetidinesulfonic acid is added as a solid (75 mg, see example 181) followed by triethylamine (0.05 ml) dropwise. The reaction is stirred at ambient temperature for 23 hours, filtered and the filter cake washed with dimethyl20 formamide. The filtrate was added to 20 ml 0.5 monobasic potassium phosphate (pH 4.5), the mixture washed with three 8 ml portions of ethyl acetate (discard), and 105 mg of tetrabutylammonium bisulfate (0.31 mmole) is added. The resulting mixture is extracted with dichloromethane (three 15 ml portions). The extracts are washed with 10% sodium chloride solution (two 15 ml portions), saturated sodium chloride solution (10 ml) and dried (Na^SO^). Filtration and concentration in vacuo yields (after heating at 32°C under high vacuum) 165 mg of oil; approximately 40% is the desired compound, and 60% dimethylformamide. -208Β) (ί)-trans-2-oxo-4-ph«iyl-3-I(phenylacetyl)amino]-1-azetidinesulfonic acid, potag»ium salt A solution of (1)-trans^-oxo-A-phenyl-l.I (phenylacetyl)amino]-1-azetidinesulfonic acid tetrabutylammonium salt in 1.5 ml acetone is treated with 41 mg (0.121 mole) of potassium perfluorobutanesulfonate. Dilution with 12 ml of ether affords a glass, which upon trituration with ether affords 43 mg of solid which contains about 201 of aa impurity containing a tetrabutylammonium moiety. Tha solid is dissolved in 50% agueous acetone and passed through e Dowex 50W-X2 X* ion-exchange resin (1 ml).

Removal of solvent yields a solid which is washed with acetone, hexane and dried (60°C, high vacuum). The yield of the title compound is ]5 mg.

Analysis calc'd for ci7®isN2O5S*K! C, 51.23; H, 3.80; N, 7.03; S, 8.05; K, 9.81 Found: C, 50.44; H, 4.20; N, 7.01; S, 7.59; X, 9.40 Example 183 (t)-(trans,Z)-3-[I2-Amino-4-thiazolyl)(methoxyimino) acetyl]amino]-2-oxo-4-phenyl-1-azetidinesulfonic acid, potassium salt A solution of N-hydroxybenzotriazole hydrate (.52 mg) and (Z)-2-amino-a-(methoxyimino)-4thiazoleacetic acid (69 mg) in dimethylformamide (0.3 ml) is treated with solid dicyclohexylcarbodiimide (70 mg) under argon, at ambient 51333 -209temperature. The resulting mixture is stirred for 1 hour, (£)°(trans)-3-amino-2-oxo-4-phenyl1-azetidinesulfonie acid (75 mg; see example 181) is added as & solid, followed by triethylamine dropwise (0.05 ml). The reaction is stirred at ambient temperature for 23 hours. The dimethylformamide is removed under high vacuum at 30°C, and the residue triturated with 2 ml of acetone and filtered. The filter cake is washed with additional acetone (two 3 ml portions) and potassium perfluorobutanesulfonate (86 mg) is added to the filtrate. Dilution with 10 ml of ether produces a gummy solid which is triturated, washed with acetone and hexane to yield, after drying, 82 mg of the title compound as a solid.

Analysis for C^H^NgOgS^K Calc'd: C, 40.26; H, 3.16; N, 15.65; S, 14.33; K, 8.74 Found: C, 38.60; 3, 3.19; N, 15.07; S, 13.87; K, 7.5 Example 184 (cis) -2-Oxo-4-phenyl-3-1 (phenylacetyl) arnino]-!azetidinesulfonic acid, potassium salt ' A) N-Benzylidene-2,4-dinethoxybenzylamine 12.0 g of 2,4-dimethoxybenzylamine hydrochloride is added to 100 ml of IN sodium hydroxide solution and the mixture is extracted with 125 ml of ethyl acetate. The organic layer is dried over anhydrous sodium sulfate and stripped of solvent to give S1303 -21010.2 g of 2,4-dimethoxybenzylaraine as an oil.

Thia amine ia dissolved in 150 ml of Benzene; 6.47 g of benzaldehyde and 0.6 g of £-toluenesulfonic acid monohydrate are added. The mixture is heated under reflux removing water with a Dean-Stark separator and in two hours the calculated amount of vater (1.1 ml] separates out. The mixture is cooled to room temperature. Upon further cooling the benzene solution deposits some precipitate. Benzene is removed under reduced pressure and 60 ml of petroleus ether is added to the residue. An oily layer separates out with more precipitate. Benzene (10 ml) is added to make the layers homogeneous and the remaining precipitate is filtered. The filtrate is stripped of solvent to give 14.2 g of the title compound as an oil. Β) (±)-(cis)-4-Phenyl-l-(2,4-dimethoxybenzyl)2-oxo-3-azidoazetidine α-Azidoacetic acid (1.62 g) is dissolved in 25 ml of methylene chloride under ..nitrogen.

To this solution are added 3.24 g of triethylamine and 1.02 g (4.0 mmole) of the imine N-benzylidene-2,4-dimethoxybenzylamine dissolved in 10 ml of methylene chloride. The mixture is cooled in an ice-bath and 3.36 g of trifluoroacetic anhydride is added slowly; the solution becomes dark colored. After stirring for 1 hour in an ice-bath, the mixture is warmed to room temperature and stirred an additional 15 minutes. -211The solution is then washed with water (60 ml), % NaHCOg solution (two 50 ml portions), and IN HCl solution (60 ml). The organic layer is dried over anhydrous sodium sulfate and stripped of solvent to give 1.72 g of crude product as dark gum. The gum is treated with charcoal several times and the resulting brown mixture is chromato graphed on 40 g silica gel eluting with 1:1 petroleum ether:ethyl acetate. The combined fractions yield crystal upon quick-freezing in a dry ice-acetone bath. Using this as a seed the product is recrystallized from petroleum ether-ethyl acetate to give 817 mg of the title compound as needles which melt upon warming to room temperature.

C) (t)- (cis)-4-Phenyl-2-oxo-3-azidoazetidine (-)-(cis)-4-Pheny1-1-(2,4-dimethoxybenzyl)2-oxo-3-azidoa2etidine (737 mg) is dissolved in ml of acetonitrile and heated to 80°-83°C under nitrogen. To the resulting solution are added over a 1 hour period 943 mg of potassium persulfate and 570 mg of potassium monohydrogen phosphate, both dissolved in 25 ml of water.

After the addition, the mixture is further heated at 80°-83°C for 7 hours. The mixutre is cooled and the pH is adjusted to 6-7 by adding solid potassium monohydrogen phosphate. Most of the acetonitrile is removed under reduced pressure and the resulting mixture is extracted with -21260 ml of chloroform. The chloroform layer is washed with water (60 ml), dried over anhydrous sodium sulfate and stripped of solvent to give a crude product as an oil. The crude product is chromatogrpahed on 40 g of silica gel eluting with 1:1 petroleum ether-ethyl acetate. The combined fractions yield crystals and the product is reerystallized from petroleum etherethyl acetate to give 267 mg of the title compound.

D) (t) - tcis) -4-Phenyl-2-oxo-3-azido—1-azetidinesulfonic acid, tetrabutyl ammonium salt (±) - (cis)-4-Phenyl-2-oxo-3-azidoazetidine (162 mg) is cooled to 0°C under nitrogen and 3.5 ml ofca£.5M dimethylformamide-sulfur trioxide complex solution in dimethylformamide is added dropwise via syringe. The resulting clear solution is stirred at 0°C for 15 minutes. The mixture is then poured into 50 ml of 0.5M monobasic potassium phosphate solution and washed with methylene chloride (three 50 ml.portions). tetra-n-Butylammonium bisulfate (292 mg) is added to the agueous solution and the mixture is extracted with methylene chloride (six 50 ml portions). The combined methylene chloride layers are dried over anhydrous sodium sulfate and stripped of solvent to give 272 mg of the title compound as a gum. 513θ3 -213Ε) (ί) - (cis) =2-0x0-4-phenyl~3-I (phenylacetyl)amino] l-azetidinesulfonic acid, potassium salt Ct)-(cis) -4 -Pbenyl-2-oxo-3-azido-l-azetidinesuIfonic acid, tetrabutyl ammonium salt (233 mg) is dissolved in 4 ml of ethanol and hydrogenated with 80 mg of platinum oxide catalyst at one atmosphere. After 1 hour stirring the catalyst is filtered through a Millipore (Trade Mark) filter with Celite; some catalyst particles pass through, the filter to give a Black filtrate. Ethanol is removed under reduced pressure and the residue is dissolved in 4 ml of dimethylformamide. N-Hydroxybensotriazole monohydrate (81 mg), mg of phenylacetic acid, and 117 mg of dicyclohexylcarbodiimide are added and the mixture is stirred for about 16 hours under nitrogen. The slurry is evaporated in vacuo and triturated with 10 ml of acetone. The resulting slurry is filtered through a Millipore filter with a Celite top and the brown filtrate is treated with 193 mg of potassium perfluorobutane sulfonate. Upon adding 20 ml of ether a gum separates out. Liquid is removed and the gum is washed with ether. The gum is dissolved in 10 ml of methanol. Upon adding ether, a small amount of precipitate is formed. The mixture is filtered and the colored filtrate is treated with more ether. The precipitate formed is filtered and recrystallized twice from ether-methanol to give 26 mg of the title compound.

Analysis calc'd for C^H^gOj-NjSK^HjO: C, 46.99; H, 4.41; N, 6.45 Found: C, 47.24; H, 4.19; N, 6.34 51333 -214gxamplc 185 (cis,2)-3-11 (2-Amino-4—thiazolyl) (methoxyimino)acetyl]amino)-2-oxo-4-phenyl-l-azetidinesulfonic acid, potassium aalt (cis)-2-Oxo-4-phenyl-3-I(phenylacetyl)amino]1-azetidinesulfonic acid, potassium salt (560 mg; see example 184, part D] is dissolved in 5 al of ethanol and hydrogenated with 110 mg of platinum oxide catalyst at one atmosphere. After one hour stirring the catalyst is filtered through a Millipore filter with Celite: catalyst particles pass through the filter to give a black filtrate. Ethanol is removed under reduced pressure and the residue is dissolved in 4 ml of dimethyl15 formamide. N-Hydroxybenzotriazole monohydrate (168 mg), 221 mg of the (2)-2-amino-a-(methoxyimino)4-thiazoleacetic acid and 227 mg of dicyclohexylcarbodiimide are added and the mixture is stirred for about 16 hours under nitrogen. The slurry Is evaporated in vacuo and triturated with 15 ml of acetone. The resulting slurry is filtered through a Millipore with Celite and the filtrate is treated with 372 mg of potassium perfluorobutane sulfonate. Upon adding 15 ml of ether a gum separates out. Liquid is removed and the gum is washed with ether. The gum is dissolved in 5 ml of water and applied on 150 ml of HP-20 resin eluting with water. Fractions (30 ml each) 16-34 are combined and lyophilized to give 201 mg of the title compound as a solid.

Analysis calc'd for CjgH^OgNjSjK·! 1/2 HjO: C, 36.73; H, 3.49; N, 14.28; S, 13.07; K, 7.97 Found: C, 36.65; H, 3.00; N, 13.99; S, 13.48; X, 8.30 -215Sxample 186 (cis)-3-Amlno°2-oxo-4-(2-phenylethenyl)-1azetidinesulfonic acid A) N-(3-phenyl-2propenylidene)-4-methoxyaniline p-Anisidine C12.32 g) is dissolved in 160 ml of methylene chloride and 20 g of anhydrous magnesium sulfate is added. The mixture is cooled in an ice hath and 13.22 g of trans10 cinnamaldehyde is added. The mixture is stirred under nitrogen for 2 hours and then filtered.

The filtrate is evaporated to give a solid.

The crude product is recrystailized from methylene chloride-petroleum ether to give 20.96 y of the title compound as a solid.

B) (i)-(cis)-3-Azido-l-(4-Methoxyphenyl)-2-oxo4-(2-phenylethenyl)azetidine 2-Azidoacetic acid (24.26 g) is dissolved 7° ih 100 ml of methylene chloride and cooled in an ice bath. To this solution is added 48.57 g of triethylamine and 14.24 g of N-f3-phenyl-2propenylidene)-4-methoxyaniline.dissolved in 250 ml of methylene chloride. To the resulting solution is added 50.41 g of trifluoroacetic anhydride dropwise over a one hour period. After stirring for one hour in an ice bath, the mixture is warmed to room temperature and stirred for about 16 hours. The mixture is then diluted with 250 ml of methylene chloride and washed 513 ί) 3 -216with water (750 ml), 5« «odium bicarbonate solution (two 75Q ml portions), and IN HCl solution (750 al). The organic layer is dried over anhydrous sodium sulfate and stripped of solvent to give a solid. The crude product is recrystallized frcm ethyl acetate to give 11.39 g of the title compound as s solid. d (i) - (cis )-3-Azido-2-oxo-4—(2-phenylethenyl) azetidine To a solution of 10.22 g ot ceric ammonium nitrate is 13 ml of water at 0°C is added 1.99 g of (ί)-(cis)-3-azido-l-(4-methoxyphenyl)-2-oxo4-(2-phenylethenyl)azetidine is dissolved in 65 ml of acetonitrile during a 15 minute period (additional 10 ml of acetonitrile is used for rinse). The mixture is stirred for an additional 15 minutes at 0°C, diluted with 750 ml of ethyl acetate, washed with water (six 600 ml portions), dried over anhydrous sodium sulfate, and stripped of solvent to give an oil. The crude product is chronatographed on 90 g of silica gel, eluting first with 250 ml of 30% ethyl acetate/petroleum ether, then 50% ethyl acetate/petroleum ether. Fractions (50 ml each) 11-16 are combined and evaporated to give 602 mg of the title compound as an oil. 5x333 -217D) (ί)—(cis)=3-Azido-2—oxo—4—(2—phenylethenyl)-1azetidinesulfonic acid, tetra-n-butylammonium salt (A) - (.cis) -3-Azido-2-oxo-4- (2-phenylethenyl) azetidine (334 mg) is dissolved in 3 ml of dimethylformamide and 868 mg of pyridine-sulfur trioxide is added. The mixture is stirred at room temperature for 40 hours under nitrogen and then poured into 200 ml of 0.5 M monobasic potassium phosphate solution and washed with 30 ml of 10 methylene ehl©ride. tetra-n-Butyl ammonium bisulfate (530 mg) is added to the aqueous solution and the mixture is extracted with methylene chloride (four 50 ml portions). The combined organic layers are back-washed with water (two 100 ml portions), dried over anhydrous magnesium sulfate and stripped of solvent to give 824 mg of the title compound as a gum.

E) (±)-(cis)-3-Azido-2-oxo-4-(2-phenylethenyl)-120 azetidinesulfonic acid (±)-(cis)-3-Azido-2-oxo-4-(2-phenylethenyl)1-azetidinesulfonic acid, tetra-n-butylammonium salt (300 mg) is dissolved in 4 ml of tetrahydrofuran and stirred rapidly. To the mixture is added 600 mg of zinc dust followed by 0.8 ml of IN monobasic potassium phosphate solution. The mixture is heated to 45°C and stirred at this temperature for 3 hours. The mixture is then filtered and the filtrate is taken in 40 ml of 2° methylene chloride and 10 ml of water. The -218agueous layer ia further extracted with methylene chloride (three 40 ml portions) and the combined methylene chloride layers are stripped of eolvent to give 256 mg of a foam. This crude product is dissolved in a small amount of ea. 301 acetone/water and applied on 7.5 ml of Dowex (X+) resin (0.7 meg./vnl) eluting vith 40 ml of water. The eluent is evaporated to give 151 mg of foam, which is dissolved in 2 ml of water and acidified to pB 2 vith IN BC1 solution.

A small amount of acetonitrile is added to dissolve the precipitate and the resulting solution is applied on 15 ml of HP-20 resin, eluting with 150 ml of water, then 101 acetone/water. Fractions (15 ml each) 2-13 are combined and evaporated to give 101 mg of the title compound as a foam.

Example 187 (t)-(cis,Z)-3-([(2-Amino-4-thiazolyl) (methoxyimino)acetyl) amino]-2-ΟΧΟ-4- (2-phenylethenyl)-1-azetidinesulfonic acid, potassium salt A solution of 68 mg of (Z)-2-amino-a(methoxyimino)-4-thiazoleacetic acid and 51 mg of N-hydroxybenzotriazole monohydrate in 2 ml of dimethylformamide is treated with 69 mg of dicyclohexylcarbodiimide. The mixture is stirred at room temperature for 30 minutes under nitrogen, (cis)-3-Amino-2-oxo-4-(2-phenylethenyl)-1azetidinesulfonic acid (90 mg; see example 186) and 34 mg of triethylamine are added and the mixture is stirred for 20 hours under nitrogen. 513 3 3 -219The Blurry is evaporated in vacuo and triturated with 10 ml of acetone. The slurry is filtered and the filtrate is treated with 113 mg of potassium perfluorobutanesulfonate. Dilution with ml of ether and filtration gives 169 mg of a solid product, which is dissolved in a small amount of ca. 10« acetonitrile/water and applied on 34 ml of HP-20 resin, eluting with 150 ml of water, then 1Q« acetone/water. Fractions (15 al each) 16-19 are combined and stripped of solvent to give 110 mg of the title compound as a solid.

Analysis calc’d for C^H^OgNgSjK-HjO: C, 40.23; H, 3.57; N, 13.80; S, 12.63; K, 7.70 Found: C, 40.03; H, 3.05; N, 13.61; S, 12.31; Κ,’ 7.56 Example 188 (cis)-3-Amino-4-(methoxycarbonyl)-2-oxo-l20 azetidinesulfonic acid A) I(4-Methoxyphenyl)imino]acetic acid, methyl ester A dry 3-necked, 1 liter flask equipped with a nitrogen inlet and stirring bar is charged with 56.88 g of MgSO^ followed by a solution of 'recrystallized anisidine (19.43 g) in dichloromethane (250 ml). After cooling to 0°C a solution of methyl glyoxylate hemiacetal (19.92 g) in dichloromethane (250 ml) is added over 1.5 hours.

After stirring an additional 20 minutes at 0°C, 51303 -220the reaction mixture is suction filtered, dried over eodiua sulfate, filtered and concentrated in vacuo to one-quarter volume. Hexane (300 ml) is added, and the solution is concentrated to an oil which semi-solidifies on standing under high vacuum at 5°C.

B) (cis) -3- (1,3-Pihydro-l, 3-dioxo-2H-isoindol2-yl) -4-methoxyearbony 1-2-oxo-1- C4-methoxyphenyl) azetidine λ dry 3-necked 500 ml flask equipped with stirring bar, addition funnel, septum and nitrogen inlet is charged with a solution of [(4-Methoxyphenyl)imino)acetic acid, methyl ester (21.09 g) in dichloromethane (150 ml) and cooled to 0°C. Triethylamine (19.2 ml) 0.14 mole is added dropwise followed by a solution of (N-phthalimido)acetyl acid chloride (28.4 g) in dichloromethane (150 ml) over 1 hour. The resulting mixture is stirred for 1.5 hours at 0°C, and diluted with 2.5 1 of dichloromethane. The organic solution is washed with pH 4.5 monobasic potassium phosphate (two 500 ml portions), 5* sodium bicarbonate (two 500 ml portions), saturated sodium chloride solution (500 ml), and dried over sodium sulfate. Filtration and concentration in vacuo yields a solid which is washed with ethyl acetate, cold acetone and hexane to yield 18.65 g of product. 221C) (cis) -4- (Me thoxy carbonyl) -1- (4 -methoxyphenyl) 2-OXO-3-H(phenylmethoxy)carbonyl]amino]azetidine A dry 500 ml flask equipped with nitrogen inlet, stirring bar, and septum was charged with 18.65 g of (cisI-3-(1,3-dioxo-2H-isoindol-2-vl)4-methoxycarbonyl-2-oxo-l-(4-methoxyphenyl)azetidine and 325 ml of dichloromethane. The resulting suspension ia cooled to -30°C, and methyl hydrazine (3.52 al] is added dropwise. The reaction is warmed fo 0°C and stirred for 1 hour.

An additional 0.4 ml of methyl hydrazine is added and the mixture is stirred for 10 minutes. This sequence is repeated with a total of 2.3 equivalents of methyl hydrazine (7.7 ml) has been added.

The solvent was removed in vacuo; 200 ml of fresh dichloromethane is added, and the mixture is again concentrated. This sequence was repeated two additional times, the resulting foam was dried under high volume for 20 minutes, redissolved in 225 ml dichloromethane, and allowed to stand at ambient temperature for about 16 hours during which time a considerable amount of solid precipitates. The mixture is filtered under nitrogen, the filtrate cooled to 0°C (nitrogen atmosphere) and treated with diisopropylethyl amine (17 ml) followed by benzyl chloroformate (7 ml) dropwise. The reaction is stirred at 0°C for 30 minutes, then at ambient temperature for 1.5 hours. The mixture is washed with two 300 ml portions of pH 4.5 monobasic potassium phosphate buffer, 5% sodium bicarbonate (two 300 ml portions), saturated sodium chloride (300 ml), -222dried (sodium sulfate), and filtered. Concentration in vacuo, yields a foam which on trituration with ether yields 9.9 g ot the title compound as a solid.

D) (cis)-4-(Methoxycarbonyl)-2-ΟΧΟ-3-ΙI(phenylmethoxy) carbonyl] amino]-1-azetldine A solution of ceric ammonium nitrate (8.59 g) in 60 al of 1:1 acetonitrile-water is treated with a slurry of 2 g (cis)-41° (ma thoxy carbony 11-1- (4-methoxyphenyll-2-oxo-3I [ (phenylmethoxy] carbonyl]amino] azetidine in 50 ml acetonitrile over 10 minutes. The reaction mixture is stirred an additional 10 minutes at ambient temperature and diluted with ethyl acetate (100 ml), lhe separated agueous layer is washed with ethyl acetate (three 40 ml portions) and the combined organic extracts are washed with 50t sodium bicarbonate (three 70 ml portions).

The basic washings are back-washed with ethyl acetate (50 ml) and the combined organic extracts are washed with agueous sodium sulfite, 5% agueous sodium carbonate (100 ml), 5t sodium chloride solution (two 100 ml portions), saturated sodium chloride (two 50 ml portions), and stirred over Darco(Trade Mark)G-60 charcoal for 30 minutes sodium sulfate is added, and the mixture is filtered and concentrated in vacuo to yield an oil which on trituration with ether yields 685 mg of the title compound as a solid. -223E) (cis)-4-(Methoxycarbonyl) -2-oxo-3-JI (phenylmethoxy) carbonyl] amino]-1-azetidinesul fonic acid, tetrabutylammonium salt A mixture of (cis)-4- (aethoxycarbonvl)-25 oxo-3-11 (phenylmethoxy) carbonyl]amino] -1-azetidine (100 mg) and 172 mg of a pyridine-sulfur trioxide complex in 1 ml of pyridine is stirred under argon for 3 hours at 80®C. The reaction mixture is poured into 70 ml of 0.5 M monobasic potassium phosphate (pH 5.5) and extracted with four 30 ml portions of dichloromethane (discard). Tetrabutylammonium hydrogen sulfate (122 mg) is added to the aqueous layer which is then extracted with dichloromethane (four 30 ml portions). The organic extracts are washed with 8% sodium chloride solution, dried over sodium sulfate and filtered. Concentration in vacuo yields 185 mg of the title compound as a viscous oil.

F) (cis)-3-Amin0-4-(methoxycarbonyl)-2-oxo-lazetidinesulfonic acid A solution of 186 mg of (cis)-4-(methoxycarbonyl)-2-oxo-3-II(phenylmethoxy)carbonyl]amino]1-azetidinesulfonic acid, tetrabutylammonium salt in 2 ml of methanol is hydrogenated over 10» palladium on charcoal (95 mg) for 1.5 hours at 1 atmosphere. The catalyst is filtered off and rinsed with dichloromethane and the filtrate is treated with 97% formic acid and cooled to 2° -50°C (the presence of a seed crystal at this stage is necessary to induce crystallization).

After crystallization commences, the mixture is allcwed to stand for about 16 hours at 10°C. -224 The resulting solid is washed with dichloromethane, hexane, and dried in vacuo, yielding 50 mg of the title compound.

Example 189 (cis)-3-II2-Amino-4-thia zolyl) I II-(diphenylmethoxycarbonyl) -1-methylethoxy] imino] acetyl] amino] -4- (methoxycarbonyl) -2-oxo-l-azetidinesulfonic acid, potassium salt λ solution of N-hydroxybenzotriazole hydrate (34 mg) and 101 mg of 2-amino-a-HI-(diphenylmethoxycarbonyl) -1-methylethoxy] imino] -4thiazoleacetic acid in 0.5 ml of dimethylformamide is treated with solid dicyclohexylcarbodiimide (45 mg) and the mixture is stirred under argon for 45 minutes (ambient temperature), (cis)-3Amino-4-(methoxycarbonyl)-2-oxo-l-azetidinesulfonic acid (45 mg; see example 188) is then added as a solid followed by triethylamine (0.03 ml) dropwise. The reaction is stirred at ambient temperature for about 16 hours. The dimethylformamide is removed under high vacuum at 30°C and the residue is triturated with acetone.

The supernatant is treated with potassium perfluorobutanesulfonate (67 mg). Dilution with ether produces a solid which is washed with ether and dried in vacuo to yield 93 mg of the title compound. -225Example 190 (cis) —3— 11 (2-iHnino"4-thiazolyl) I fl-earbaxy-lmethvlethoxy) imino]acetyl]amino]-4-(methoxycarbonyl) -2-oxo-l-azetidinesulfonic acid, dipotassium aalt A slurry of Ccis)-3·- [ I2-Amino-4-thiazolyl) III-(diphenylmethoxyearbonyll-1-methylethoxy)imino] acetyl] amino]'-4- (me th oxycarbonyl) -2-oxo-lazetidinesulfonic acid, potassium salt in 0.4 ml of anisole is stirred at -12°C under argon, and 0.9 ml of cold (-10°C) trifluoroacetic acid is added. After 1.5 hours, 4 ml of ether and 2 ml of hexane are added and the resulting slurry is stirred for 15 minutes at -10°C, then 15 minutes at ambient temperature. The solid is isolated by centrifugation and washed with ether. The pH of a suspension of this material in 0.5 ml of cold water is adjusted to 6 with IN potassium hydroxide and then applied to a 30 ml HP-20AG column. Elution with water yields 30 mg of the title compound after evaporation (acetonitrile added and evaporated twice). Analysis calc'd for C^H^I^NgOgS^: C, 31.15; H, 2.81; N, 12.98 Found: C, 29.08; H, 3.03; N, 12.19 -226Example 191 (5)-(trans)-3-Amino-4-ethynyl-2-oxo-l-azetidinesulfonic acid A) 2-(Trimethylsilyl)ethynylmagnesium bromide To a flame-dried 50 ml flask maintained under positive nitrogen pressure is added 20 ml of dry tetrahydrofuran, 2.20 ml of trimethylsilyl acetylene and 5.05 ml of 3.06 M solution of methylmagnesium bromide in ether. The mixture is stirred for 140 minutes yielding the title compound.

B) (S)-(trans)-4-J2-(trimethylsilyl)ethynyl]-2oxo-3-[(triphenylmethyl)amino)azetidine ' To a flame dried 250 ml 3-necked flask is added 6.00 g of (S)-(cis)-4-(methylsulfonyl)2-ΟΧΟ-3-Ι(triphenylmethyl)amino]azetidine. The flask is flushed with nitrogen, and then maintained under positive nitrogen pressure.

After the reaction mixture is cooled in a dry ice/isopropanol bath, 4.65 ml of a 3.06 M solution of methylmagnesium bromide in ether is added dropwise via syringe with rapid stirring. The solution of 2-(trimethylsilyl)ethynylmagnesium bromide prepared in part A is added via a Teflon(Trade Mark) tube under positive nitrogen pressure (the flask containing the reactant is rinsed with 7 ml of tetrahydrofuran). When the addition is complete the cold bath is removed. After 45 minutes, a solution of 3.5 g -227of potassium bisulfate in 20 ml of water is added. Most of the tetrahydrofuran is removed on the rotary evaporator. The residue is transferred to a separatory funnel with ether and water. The water layer is separated and extracted twice with ether. The combined ether layers are washed once with saturated aqueous sodium chloride, dried over sodium sulfate, and filtered. Removal of the solvent gives a foam which is chromatographed on a silica column. Elution with 2 liters of dichloromethane, 1 liter of 1% ether/dichloromethane, 2 liters of 2% ether/dichloromethane and 1.5 liters of 10% ether/dichloromethane (fraction 1=1000 ml; fraction 2,3=500 ml; fraction 4-end®250 ml) gives 1.30 g of the title compound in fractions 2-8 and 1.80 g of the corresponding trans-isomer in fractions 12-19. Fractions 9-11 contain 1.19 g of a mixture of cis and trans isomers.

C) (S)-(trans)-4-ethynyl-2-oxo-3-I(triphenylmethyl)amino]azetidine (S) - (trans) - 4 - [ 2- (tr ime thyl s i ly 1, e thynyl] 2-oxo-3-[(triphenylmethyl)amino]azetidine (2.97 g) is dissolved in 30 ml of dichloromethane and 330 mg of tetrabutylammonium fluoride (containing 20-25% water) is added. After 20 minutes, the solvent is removed in vacuo. The residue is taken up in ethyl acetate and water. The organic layer is separated, washed once with water and once with saturated aqueous sodium chloride, 31393 -228 " dried over sodium sulfate, and filtered.

Removal of the solvent gives an oil whieh is stirred for 15 minutes with 60 ml of pentane to afford 2.35 g of the title compound as a powder (after drying in vacuo).

D) (5)-(trans)-3-Amino-4-ethynyl-2-oxo-lazetidinesulfonic acid (5)-(trans)-4-ethynyl-2-oxo-3- [(triphenyl10 methyl)amino]azetidine (404 mg) and 560 mg of a complex of pyridine and sulfur trioxide are added to a 25 ml flask. After the flask is flushed with nitrogen, 4.0 ml of dry pyridine is added and the mixture is heated at 80-85°C for 3 hours. The mixture is added to a rapidly stirred mixture of 4.0 ml of concentrated hydrochloric acid, 50 ml of water, and 50 ml of ethyl acetate. The pH is adjusted to 3.15 with sodium carbonate. The water layer is separated and extracted once with ethyl acetate. The combined organic layer is washed once with saturated aqueous sodium chloride, dried over sodium sulfate and filtered. Solvent removal in vacuo gives a foam which is taken up in ml of dichloromethane. Formic acid (98%, 8 ml) is added, and after 15 minutes the mixture is concentrated to 4 ml and 10 ml of dichloromethane is added to give a solid suspended in solution. Filtration gives 100 ml of the title compound as a solid (obvious discoloration with melting >180°C). -229Example 192 13S— 13η (Ζ) ,4 8] ] -3-1 [ (2-A-Tiino-4-thia zolyl) (methoxyimino)acetyl) amino]-4-ethynyl-2-oxo-l-azetidinesulfonic acid, potassium salt 5 (Z)-2-Amino-a-(methoxyimino)-4-thiazoleacetic acid (100 mg), 85 mg of N-hydroxybenzotriazole monohydrate, and 113 mg of dicyclohexylcarbodiimide are weighed into a 10 ml flask. The flask is flushed with nitrogen and cooled in an ice-water bath. Then 0.6 ml of dimethylformamide is added and the mixture is stirred for 10 minutes, at which point an additional 0.6 ml of dimethylformamide is added. (S)-(trans)-3-Amino-4-ethynyl-2-oxo-l15 azetidinesulfonic acid (95 mg; see example 191) is added as a solid with 1.0 ml of dimethylformamide and 56 μΐ of triethylamine. The cold bath is removed and the mixture is stirred for 22 hours. Acetone (3 ml) is added and the solids present are removed by filtration and washed with an additional 4 ml of acetone.

All solvents are removed in vacuo and the residue is taken up in 5 ml of methanol and 162 mg of potassium perfluorobutanesulfonate is added and dissolved. After standing, a solid is deposited and then isolated by centrifugation to afford 68 mg of the title compound, melting point >230 C. 230Example 193 (S)-3-ί [ [ (2,5-Dichlorophenyl)thio]acetyl]amino32-oxo-l-azetidinesulfonic acid, potassium salt 3-Amino-2-oxo-l-azetidinesulfonic acid (100 mg) is dissolved in dry dimethylformamide (2 ml) with triethylamine (0.083 ml). 2,5Dichlorophenylthioacetic acid (123 mg) 0.502 mmol, N-hydroxybenzotriazole hydrate (81 mg) and dicyclohexylcarbodiimide (124 mg) are added, the mixture is stirred for 2 hours at room temperature, and then 2 days at 5°C. Solvent is removed in vacuo, the residue is taken up in water, filtered through Celite, and the filtrate is washed with ethyl acetate. The aqueous layer is combined with dichloromethane, tetrabutylammonium bisulfate (612 mg) is added, the pH was raised to 3 with IN potassium hydroxide solution, and after extracting a total of three times with dichloromethane, the combined extracts are dried (Na^SO^), and solvent is removed in vacuo yielding an oil. A solution of the oil in acetone is added to a solution of potassium perfluorobutanesulfonate (612 mg) in acetone causing precipitation of the product.

After addition of a small amount of ether the solid is collected by filtration, washed several times with acetone, and dried to give a powder (206 mg).

Anal. Calc'd for C^H^O^d^: C, 31.21? H, 2.14? N, 6.62; Cl. 16.75 Found: C, 27.90; H, 2.11; N, 5.84; Cl, 18.04 - 231 Exarpie 194 ( 35-trans) -3- [[1(2, 5-3ichlorophenyl) thiojacetyl] aminoj-4-methyl-2-oxo-l-azetidinesulfonic acid, potassium salt (3S-trans)-3-Amino-4-methyl-2-oxo-lazetidinesulfonic acid (250 mg; see example 139) is dissolved in dimethylformamide (2 ml) with triethylamine (193 μΐ). 2,5-Dichlorophenylthioacetic acid (285 mg), N-hydroxybenzotriazole 1θ hydrate (213 mg) and dicyclohexylcarbodiimide (287 mg) are added. After stirring for about 16 hours at room temperature, the mixture is filtered and solvent is removed in vacuo. The residue is taken up in water and filtered.

The filtrate is washed with ethyl acetate, layered with dichloromethane and tetrabutylammonium bisulfate (4.2 mmol) is added. After a total of three extractions with dichloromethane, the combined extracts are dried (NajSO^) and solvent is removed in vacuo yielding an oil (920 mg). To a solution of the oil in acetone is added potassium perfluorobutanesulfonate (946 mg) dissolved in acetone. A solid slowly precipitates, is collected, washed twice with ether, and dried to give a powder (396 mg).

Chromatography on HP-20 resin (100 ml column), eluting with 20% acetonitrile: 80% water, yields the desired product, which crystallizes upon evaporation of a water:methanol mixture.

Trituration of the residue with acetone gives a powder (233 mg); melting point 212-213°(dec.).

Anal. Calc'd for ci2HnN2°5C}-2* S2K: C, 32.95; H, 2.54; N, 6.41; Cl, 16.21; S, 14.66 Found: C, 32.91; H, 2.60; N, 6.42; Cl, 16.50; S, 13.77 - 232 Examples 195-196 Following the procedure of example 133, but substituting (3S-trans)-3-amino-4-methyl2-oxo-l-azetidinesulfonic acid for (3S-cis)5 3-amino-4-methyl-2-oxo-l-azetidinesulfonic acid and the acid listed in column I for IZ)—2— amino-ο-(methoxyimino)-4-thiazoleacetic acid, yields the compound listed in column II. 195. (R)-[(aminooxoacetyl)amino](4hydroxyphenyl)acetic acid IS [3S-(3a (R*),46]]-3-[[[ (aminooxoacetyl)amino](4-hydroxyphenyl)acetyl]amino]-4-methyl-2-oxo-lazetidinesulfonic acid, potassium salt 196. (R)-[(aminooxoacetylJamino]phenylacetic acid (3S-[3o(R*),46])-3-(([(aminooxoacetyl) amino]phenylacetyl) amino]-4methy1-2-oxo-1-azetidinesulfonic acid, potassium salt; melting point 187°C, dec. - 233 Example 197 [3S(R°)]-3-[|[ (Aminooxoacetyl lanno] (4-hycroxyphenyl) acetyl]amino]-2-oxo-l-azetidinesulfonic acid, potassium salt Following the procedure described in Example 28, but substituting (R)- I(aminooxoacetyl)amino](4-hydroxyphenyl)acetic acid for (Z)-2-aminoa-[12-(diphenylmethoxy)-1,l-dimethyl-2-oxoethoxy]imino]-4-thiazoleacetic acid, yields the title compound melting point 128°C, dec.

Example 198 [3S(R»l]-3-[[(2-ftmino-4-thiazolyl)[[[3-1(2-furanylmethylene)amino]-2-oxo-l-imidazolidinyl]carbonyl]15 amino]acetyl]amino]-2-oxo-l-azetidinesulfonic acid, potassium salt Following the procedure of example 6, but substituting (R)-2-amino-a-[[[3-I(2-furanylmethylene) amino]-2-oxo-l-imidazolidinyl]carbonyl]20 amino]-4-thiazoleacetic acid for aminothiazoleacetic acid yields the title compound, melting point >250°C. 513 9 3 - 234 Example 199 Biological Production of EM5117 9 Biter Fermentation Chromobac ter ium violaceum SC 11,378 A.T.C.C.

No. 31532 is maintained on the following sterilized agar medium (A): Crams Yeast Extract 1 Beef Extract 1 NZ Amine A 2 Glucose 10 Agar 15 Distilled HjO to 1 liter The pH is adjusted to 7.3 before sterilization at 121°C for 30 minutes.

A loopful of surface growth of the microorganism is used to inoculate each of three 500 ml Erlenmeyer flasks, each containing 100 ml of the following sterilized medium (B): Crams Oatmeal 20 Tomato Paste 20 Tap HjO to 1 liter Adjust pH to 7.0 before sterilization at 121°C for 15 minutes.

The flasks are then incubated at 25°C on a rotary shaker (300 rpm; 2 inch stroke) for approximately 24 hours.

After the appropriate incubation as described above, lt (vol/vol) transfers are made froa the growth culture flasks to one hundred 500* ml Erlenmeyer flasks each containing 100 ml of 513 9 3 - 235 the following sterilized medium (C): Oatmeal Grams 20 Tomato Paste 20 Glucose 30 Tap HjO te 1 liter The pH is adjusted to 7.0 before sterilization of 121°C for 15 minutes.

After inoculation, the flasks are incubated at °C on a rotary shaker (300 rpm; 2 inch stroke) for approximately 18-24 hours. At this time the contents of the flasks are pooled and the broth is centrifuged yielding approximately 9 liters of supernatant broth. 250 Liter Fermentation A loopful of surface growth from an agar slant (medium A) of Chremobacterium violaceum SC 11,378 A.T.C.C. No. 31532 is used to inoculate • 20 each of five -500 ml Erlenmeyer flasks each containing 100 ml of sterilized medium (B).

The flasks are then incubated at 25°C on a rotary shaker (300 rpm; 2 inch stroke) for approximately 24 hours. After the appropriate incubation, as described above, 1% (vol/vol) transfers are made from the grown culture flasks to five 4 liter Erlenmeyer flasks each containing 1.5 liters of sterilized medium B. After inoculation the flasks are then incubated at 25°C on a rotary shaker (300 rpm; 2 inch stroke) for approximately 24 hours. After the - 2.36 appropriate incubation as described above, a It transfer (vol/vol) ia made to an agitator equipped fermentation tank containing 250 liters of sterilized medium (C). After inoculation the fermentation is continued under the following Q conditions: temperature - 25 Ct pressure - 10 psig; aeration - 10 cfa; agitation - 155 rpm. Ucon Trade Mark is added as needed as antifoam agent. After approximately 18-24 hours the fermentation is completed. The fermentation broth ia then adjusted to pH 5.0 using HCl and the broth contents of the tank is centrifuged yielding approximately 230 liters of supernatant broth.

Isolation and Purification The broth supernatant from the 250 liter fermentation is adjusted to pH 5 using sulfuric acid and filtered using 3-5% diatomaceous earth (Celite). The broth filtrate is extracted with two 30 liter portions of 0.005 M cetyldimethylbenzylamraonium chloride in methylene chloride.

The combined lower phase is extracted with 6 liters of 0.05 M sodium iodide which has been adjusted to pH 5 with acetic acid. The lower phase is discarded and the upper phase is concentrated in vacuo to 500 ml.

The concentrated material is extracted with 400 ml of n-butanol. The upper phase is S i 3 3 3 - 237 discarded and the lower phase is concentrated to dryness in vacuo. The residue is dissolved (to the extent possible) in 150 ml of methanol.

The insoluble material is discarded and the methanol solution is concentrated to dryness in vacuo, yielding 38.6 g of crude antibiotic.

The crude product is dissolved in 10 ml of methanol-water (1:1) and chromatographed on a 500 ml column of cross-linked dextran gel (Sephadex G-1Q) in the same solvent mixture, eluting at 2 ml/minute and collecting 20 ml fractions. Active fractions (19-26) are combined and concentrated in vacuo. The residue (5.23 g) is mixed with 50 ml of methanol. Insoluble material is filtered out and discarded. The filtrate is concentrated in vacuo.

The residue, 5.0 g of material, is dissolved in 10 ml of pH 5 sodium 0.01 M phosphate buffer and applied to a column of DEAE cellulose (Whatman Trade Mark DE 52 Cellulose) packed and equilibrated in the same buffer. The column is eluted at 5 ml/minute with a linear gradient prepared from 4 liters of pH 5 sodium 0.01 H phosphate buffer and 4 liters of pH 5 sodium 0.1 M phosphate buffer, collecting 20 ml fractions. Active fractions (192-222) are combined and concentrated in vacuo, and methanolinsoluble material is removed, washing well with methanol. Removal of solvent leaves 576 mg of material. - 238 The 576 mg of residue is dissolved in 4 ml of water and the pH adjusted to 5 with about 1 ml of 0.1 N sodium hydroxide. The solution is chromatographed on a column of alkylated crosslinked dextran gel (Sephadex LH-20) in water, eluting at 1 ml/minute and collecting 10 ml fractions. Active fractions (38-44) are combined and concentrated, giving 459 mg of residue.

Three hundred and forty eight (348) mg of the above residue is dissolved in water and the solution placed on a column of macroreticular styrene-divinylbenzene copolymer resin (Diaion HP20AG); the column has been first prepared by washing with methanolic potassium hydroxide, methanol, methanolic hydrogen chloride, methanol and water, and then packed in water. The column is eluted with water at 1 ml/minute, collecting 10 ml fractions.

Active fractions (36-43) are combined and concentrated giving 186.4 mg of material. Chromatography (in the same way) of another 100 mg of the 459 mg of residue from the previous step yields 51.5 mg of material. At this stage the 186.4 mg of material and 51.5 mg of material are nearly pure EM5117 as shown by thin-layer chromatography and nuclear magnetic resonance spectra.

The 186.4 mg portion of EM5117 from above is dissolved in water and passed through a column of ion-exchange resin (Dowex 50W-X2, 100-200 mesh, potassium form), washing with two bed volumes of water. Concentration of the effluent - 239 yields 189.0 mg of crystalline solid. Recrystallization of this material is accomplished by dissolving it in 0.38 ml of water and adding 3.42 ml of methanol. The resulting mixture is cooled on ice and filtered, yielding 145 mg of crystals.

Two further recrystallizations in this manner from water-methanol, 1:9, yields 95.9 mg of EM5117, potassium salt, m.p. 194°(dec.).

Optical rotation in water at 21°C (c»l) 1 (mn) la] 589 +94.3° 579 +98.6 54 6 +113.1 15 436 +203 365 +34 8 The following fermentation media are effective for the production of EM5117, and may be-substituted for medium (B) and (C) in the above example. 20 MEDIUM D Grams Hutrisoy Flour 30 Glucose 50 Yeastamine 2.5 25 CaCO3 7 Distilled H^O to 1 liter MEDIUM E Grams Yeast Extract 4 30 Malt Extract 10 Glucose 30 Glycerol 2 Distilled H^O to 1 liter Adjust pH to 7.3 before sterilization MEDIUM Ρ Glycerol L-asparagine xh2po4 Na2HPO4 Glucose Grams 10 5 1 2 50 MgSO4.7H2O Yeast Extract 0.2 2.5 Tap HjO to 1 liter MEDIUM G Grams (nh4,2so4 L-asparagine Glucose 2 5 50 Glycerol 10 kh2po4 k2hpo4 MgSO4.7H2O Yeast Extract 3 7 0.2 2.5 Distilled HjO to 1 liter Adjust pH to 7.0 MEDIUM H Grams K2HP°4 kh2po4 Na Citrate 7 3 0.5 MgSO4 (NH4)2s°4 Glucose 0.1 1 30 Yeast Extract 2.5 Distilled HjO to 1 liter - 241 MEDIUM I Grams Nutrisoy Flour 10 2SO4 5 Glucose 50 Yeast Extract 2.

CaCOg 5 Distilled HgO to 1 liter MEDIUM J Grams Gerber’s Baby Oatmeal 20 Contadina Tomato Paste 5 Glucose 20 Tap HgO to 1 liter Adjust pH 7.0 MEDIUM K Grams Gerber’s Baby Oatmeal 5 Contadina Tomato Paste 20 Glucose 20 Tap HgO to 1 liter Adjust pH 7.0 MEDIUM L Grams Yeast Extract 4 Malt Extract 10 Glucose 34 MEDIUM M Grams Amberex (1003) 5 Glucose 30 Tap HgO to 1 liter - 242 MEDIUM Μ Amberex Cerelose Trade Mark 5 Tap HjO to 1 liter Grams 513 3 3 243 10 Yeast Extract Beef Extract NZ amine A Glucose Agar Distilled H^O to Example 200 Biological Production of EM5210 Gluconobacter species SC11,435 A.T.C.C.

No. 31581 is maintained on the following sterilized agar medium (A): Grams 1 1 2 liter The pH is adjusted to 7.3 before sterilization at 121°C for 30 minutes.

A loopful of surface growth from the agar slant (medium A) of Gluconobacter species SC11,435 is used to inoculate each of three 500 ml Erlenmeyer flasks each containing 100 ml of the following sterilized medium (B): Grams Yeast Extract 4 Malt Extract 10 Dextrose 4 Distilled H20 1 liter The pH is adjusted to 7.3 before sterilization at 121°C for 15 minutes.

After inoculation, the flasks are incubated at 25°C on a rotary shaker (300 rpm; 2 inch stroke) for approximately 24 hours. After the appropriate incubation, as described above, 1% (vol/vol) transfers are made from the grown culture flasks to one hundred 500 ml Erlenmeyer flasks each containing 100 ml of the following - 244 sterilized medium (C): Grams Yeast Extract 5 Glucose 10 Distilled HjO 1 liter The medium is sterilized at 121°C for 15 minutes.

After inoculation, the flasks are incubated at 25°C on a rotary shaker (300 rpm: 2 inch stroke) for 18 hours. At this time the contents of the flasks are pooled and the broth is centrifuged yielding approximately 9 liters of supernatant broth.

Isolation and Purification (small scale) Activity from the broth supernatant (10 liters) is absorbed on a 500 g column of strongly basic anion exchange resin with quaternary ammonium groups attached to a styrene-divinylbenzene copolymer lattice (Dowex AG1-X2(Cl)), washed with water and eluted with 5% sodium chloride in aqueous 0.01 M NaH^PO^. The eluate is concentrated in vacuo.

The residue is adsorbed on 250 g of charcoal which is washed with water. EM5210 is eluted with methanol-water (1:1). The active fractions are combined and concentrated in vacuo yielding crude EM5210.

The crude EM5210 is chromatographed on a 280 ml column of strong base anion exchange resin (Bio.Rad Trade Mark AG 1-X2 (C1 )) using a linear gradient prepared from 1 liter of water and 5i393 - 245 1 liter of 2M pyridinium acetate (pH 4.5). The active fractions are combined and concentrated in vacuo.

The partially purified EM5210 is further purified by gel filtration of the residue on a 500 ml column of cross-linked dextran gel (Sephadex G-10) eluting with water. The active fractions are combined and concentrated yielding 26 mg of EM5210. The potassium salt of.EM5210 is prepared by passing EM5210 through a column of cation-exchange resin (Dowex 50-X2) in the potassium form.

Isolation and Purification (large scale) The broth filtrate of a 250 liter fermentation (pH 3.7) of Gluconobacter species SC11,435 is absorbed on 10.8 kg of strong base anion exchange resin (Dowex 1-X8(Cl")). The resin is washed with water and eluted with 5% sodium chloride in 0.01 M sodium dihydrogen phosphate. The active fractions are combined and concentrated to a small volume. The precipitated salt is removed and the filtrate is desalted by passing it through a column of charcoal (1.1 kg of 20-40 mesh Darco) in water. The column is washed with water and the EM5210 is eluted with methanolwater, 1:1. Active fractions are combined and concentrated. The residue (16 g) is dissolved in water and chromatographed on a column (600 ml) of strong base ion exchange resin (Bio.Rad AG 1X2(C1~), 200-400 mesh), eluting with a linear - 246 gradient prepared iron 1 liter of water and 1 liter of 10« «odium chloride in 0.01 M sodium dihydrogen phosphate. Active fractions are combined, concentrated in vacuo to a small volume, and precipitated salts are removed by filtration.

The filtrate is applied to a column of macroreticular styrene-divinylbenzene copolymer resin. The imm ig eluted with water. The active fractions are combined, concentrated to a small volume and freeze-dried, yielding 120 ag of the sodium salt of EM5210.

To transform the sodium salt to the lithium salt an ion-exchange resin (Dowex 50 W-X2, lithium form) coluan is used. The sodium salt (100 mg) is dissolved in 0.5 ml of water, applied to the oolum and eluted with water. The active fractions are combined and directly freeze-dried, yielding 95 mg of the lithiun salt of ΣΜ5210 or as an anorphous solid.

The free acid (inner salt) of EM5210 is prepared by passing a base salt of EM5210 through a column of weak acid ion-exchange resin in the H* form. For example, about 2.5 mg of the lithium- salt can be applied to a column of Bio.Rad Bio.Rex 70 (H+) and eluted with water to give 1.45mg of the free acid (inner salt).

Chemical properties of EM5210 1) Ninhydrin positive. 2) Acid hydrolysis (6N HCl at 115°C for 16 hours) gives two major ninhydrin positive spots by paper chromatography (Whatman No. 1, butanol-acetic - 247 acid-water (5:1:4), and one weak ninhydrin positive spot that quenches UV-excited fluorescence. The two major ninhydrin positive spots are D-glutamic acid and D-alanine.

Physical characteristics of EM5210 1) UV spectrum of the sodium salt in water: end absorption 2) IR - Major peaks of the lithium salt in KBr: 1770, 1640, 1530, 1384, 1242, and 1051 caTl. 3) PMR - Chemical shifts of the lithium salt ia deuterated water, ppm down field from TSP: 1.40 (d, J-7Hz), ea. 2.14 (m), ca. 2.44 (m), 3.49 (s) 3.73 (t,J=6Hz), 3.94 (s), 4.28 (m) Optical rotation of the free acid (inner salt) in water at 24°C (C^O.15%) (pH 2.7): i (nm) Ial 589 +73° 578 +79° 54 6 +91° 436 +159° 365 +263° The following fermentation media have been found effective for the production of EM5210 and may be substituted for medium (B) and (C) in the text.

MEDIUM D Oatmeal Tomato Paste Grams - 248 MEDIUM D (continued) Gram» Tap HgO to Adjust pH to 7.0 121°C for 15 minutes. 1 liter before sterilization MEDIUM E Grams Yeastamine 5 Cerelose 11 Tap HgO to 1 liter Sterilization at 121°C for 15 minutes. MEDIUM F Grams Glucose 5 Tartaric Acid 2 Yeast Extract 0.5 (nh4)2po4 1 (NH4)2SO4 2 KgHPO4 0.5 NaHgPO4 0.5 MgSO4.7HgO 0.2 CaCOg 1 Distilled HgO 1 liter Adjust pH to 6.0 before sterilization 121°C for 15 minutes. MEDIUM G Grams Nutrisoy flour 30 Glucose 50 Yeastamine 2.5 CaCOg 7 Distilled HgO to 1 liter MEDIUM G (continued) 121°C for 30 minutes Sterilization at MEDIUM H 5 Grams NZ Amine A 10 Cerelose 33 Yeastamine 2.5 Tap HjO to 1 liter XO Sterilization at 121°C for 15 minutes MEDIUM I Grams Nutrisoy flour 15 15 Soluble starch 15 Glucose •50 CoC12-6H2O 0.005 CaCo3 10 Distilled H2O to 1 liter 20 Sterilization at 121°C for 30 minutes - 250 Biological Activity The following methodology is used to determine the minimum inhibitory concentration (hereinafter referred to as MIC) of the e-laetams of this invention.

The test organisms are grown in approximately 15-20 al of Antibiotic Assay broth (Difco) Trade Mark by inoculating (in tubes) the broth with a loopful of the organism from a BHI (Difco) .agar slant.

The inoculated tubes are incubated at 37°C for to 20 hours. These cultures are assumed to g contain 10 eolony forming units (hereinafter CPU) per milliliter. The cultures are diluted 1:100 to give a final inoculum level of 10* CPU; dilutions are made with X-10 broth*.

The compounds are dissolved in the appropriate diluent at a concentration of 1000 ng/ml. Twofold dilutions are made in X-10 broth resulting in a range from 1000 ng/ml to 0.5 ng/ml. 1.5 ml of each dilution is placed into individual square petri dishes to which 13.5 ml of K-10 agar** is added. The final drug concentration in the agar ranges from 100 ng/ml to 0.05 ng/ml. Organism growth control plates containing agar only are prepared and inoculated before and after the test plates. The organisms are applied to the agar surface of each plate with the Denley Multipoint Inoculator (which delivers approximately 0.001 ml of each organism) resulting in a final a inoculum level of 10 CPU on the agar surface. 51333 - 251 The plates are incubated at 37°C for IS hours and the MIC's are determined. The MIC is the lowest concentration of compound inhibiting growth of the organism.

•K-10 broth is a yeast beef broth containing: Beef extract 1.5 g Yeast extract 3.0 g Peptone 6.0 g Dextrose 1.0 g Distilled water q.s. 1 liter agar Beef extract 1.5 g Yeast extract 3.0 g Peptone 6.0 g Dextrose 1.0 g Agar 15.0 g Distilled water q.s. 1 liter The tables that follow are tabulated results obtained when the β-lactams of this invention are tested against various organisms. The number following each organism refers to the number of the organism in the collection of E. R. Squibb & Sons, Xnc., Princeton, New Jersey. A dash (-) in the tables means that the compound tested did not show activity against the particular organism at 100 ug/ml. The symbol N.T. means not tested.

Claims

1. A compound of the formula:- wherein R-j is hydrogen or acyl or the group Rj-HH- is protected amino; R^ and are the same or different and each -is hydrogen, alkyl, cyeloalkyl, phenyl or substituted phenyl, or one of R^ and R^ is hydrogen and the other is alkoxycarbonyl, alken-1-yl, aliynl-yl, 2-phenylethenyl or 2-phenylethynyl; V is a leaving group; and M + is hydrogen or a cation.

2. A compound of the formula :ΔΜΗCH —KHSOj M wherein A is a protecting group and V, R^, R^ claim 1. are as defined in 513 9

3.- 272 A process for preparing a compound of the formula:- HHSOj M wherein A is a protecting group and V, Ry R^ and M are as defined in clain 1 which comprises sulfonating a conpound of the foroula:- wherein the symbols are as defined above. - 273

4. A compound according to Claim 2, wherein one of R^ and R^ is hydrogen and the other is alkyl.

5. A compound according to Claim 4, wherein one of R_ + ri and R^ is hydrogen and the other is methyl and M is hydrogen.

6. A compound according to any one of Claims 1,2,4 and 5 wherein V is OMs.

7. A process according to Claim 3, wherein V is OMs.