2-(2-SUBSTITUTED PYRROLIDIN-4-YL)THIO-CARBAPENEM DERIVATIVES

TECHNICAL FIELD

The present invention relates to a novel carb'apenem derivative useful as a therapeutic agent for treatment of bacterial infectious diseases in pharmacological field.

BACKGROUND ART

Recently, beta-lactam antibiotics having a beta-lactam ring in their structurs as in penicillin derivaties have been discovered in the natural world. As the typical exam¬ ple thereof, thienamycin having the following structural formula was first isolated by fermentation of microorganism Streptomyces cattleya (Journal of American. Assoc. Vol. 100, p6491, 1978) .

Thienamycin = [5R-[5oC , 6 o. (R ) ] ]-3-[ (2-aminoethyl) - thio]-6-(1-hydroxyethyl) -7-oxo-l-azabicyclo[3.2.0]- hept-2-ene-2-carboxylic acid

According to the result of antimicrobial activity test, it has been identified that thienamycin exhibits a ^' broad and

potent antimicrobial activity against gram-positive and gram-negative bacteria. Thus, thienamycin was expected as a beta-lactam antibiotic substance having a high clinical utility. However, it has been reported that since thiena- ycin itself is chemically unstable arid can be readily decomposed in vivo by dehydrogenase-l(DHP-I enzyme) present in kidney, when thienamycin is clinically administered, antibacterial activity in vivo exhibits a tendency to reduce and the recovery rate in urine is extremely low (Antimicro. Agent. Che other. Vol 22, p62, 1982) . Thus, in order to prepare the compound having an improved chemical stability while maintaining a good -antibacterial activity of thienamy¬ cin numerous thienamycin derivatives have been synthesized. Among such thienamycin derivatives, particularly imipenem, i.e. (5R, 6S, 8R) -3-[ [2- (f ormimidoylamino) ethyl] thio] -6- (1- hydroxyethyl) -7-oxo-l-azabicyclo[ 3.2.0]hept-ene-2-carboxylic acid hydrate which is prepared by formylating an amino group in thienamycin, exhibits antibacterial activity equivalent to, or higher than, that of thienamycin against various bacteria including beta-lactamase producing strains and particularly a potent antibacterial activity against Pseudo- monas aeruginosa, which is 2 to 4 times stronger than that of thienamycin and further shows a slightly improved stability in a solution in comparison withy thienamycin. Accordingly, imipenem was developed as a pharmaceutical medicament which can be practically and clinically utilized (J. Med. Chem. Vol. 22, pl435, 1979) . However, since imipe¬ nem can be readily decomposed by DHP-I enzyme present in human kidney as like as thienamycin, it cannot be used for treatment of urinary tract infection and further, substances produced by DHP-I enzyme decomposition can induce a serious renal toxicity. Accordingly, imipenem cannot be adminis¬ tered alone and should be administered together with DHP-I enzyme inhibitors such as cilastatin (J. Antimicrob. Chemo. Vol. 12 (Suppl. D) pl(1983)). Moreover, recently a frequent use of imipenem for prophylaxis and treatment of infectious diseases results in remarkable increase of i ipenem-resistant Straphylococcus aureus and Pseudo onas aeruginosa strains in

clinical field. Imipenem cannot provide a suitable thera¬ peutic effect on diseases caused by such resistant strains.

As a result of an effort to solve such disadvantages, many antibiotics having chemical structure and pharmacologi¬ cal activity similar to imipenem but not having the above- mentioned disadvantages involved in imipenem have been developed. For example, they are disclosed in European Patent No. 411664A, 272456, 272457, 280771, 341557 and the like. Among these patent specifications, European Patent No. 411664A discloses a carbapenem compound having 2-[2- (aminocarbonyl)vinyl]pyrroldin-4-yl]-thio group at 2-posi- tion of a carbapenem basic structure with a specific example of (1R, 5S, 6S)-2-[(2S, 4S)-2-[ (E)-2-(aminocarbonyl)vinyl]- pyrrolidin-4-yl-thio]-6-[ (R)-1-hydroxyethyl]-1-methyl-l-car- bapen-2-em-3-carboxylic acid (Bo-2171, compound of Example 3) . Such beta-lactam antibiotics generally exhibit a toxic effect selectively only on pathogenic bacteria with substan¬ tially no toxic effect on animal cells. Accordingly, they have been broadly and safely used for treatment of infec¬ tious diseases caused by bacteria for clinical purpose. However, since these beta-lactam antibiotics do not suffi¬ ciently exhibit a satisfactory antibacterial effect on causative microorganisms for incurable infectious disease, such as Staphylococcus aureus and Pseudomonas aeruginosa which are resistant to methicillin, their clinical use is greatly restricted, particularly in immunodeficient patients from which such resistant strains are frequently isolated. Further, although such known antibiotic compounds have a resistance to DHP-I enzyme to some extent, it is not suffi¬ cient to the desired extent. Accordingly, the development of antibiotic compounds showing an improved antibacterial activity against such resistant strains has been continuous¬ ly required. Specifically, the major aspect of development of a novel carbapenem 'antibiotic compound resides in an increase of resistance against DHP-I enzyme and a reduction of renal toxicity and side effects on central nervous system as well as an increase of antibacterial activity.

DISCLOSURE OF INVENTION

Thus, the present inventors have conducted an extensive study to provide a novel carbapenem compound showing an excellent antibacterial activity and a strong resistance against DHP-I enzyme. As a result, we have found that a novel group of carbapenem compounds having a moiety of the following formula(A) at 2-position of carbapenem structure satisfies the above-mentioned requirement and then completed the present invention :

wherein R3 represents hydrogen or lower alkanimidoyl,

X II Z represents -C-R ₄ or R _g ,

X represents 0 or NH,

R ₄ represents amino or heterocyclic amine group, each of which can be unsubstituted or substituted with a group

of formula -CH_ , a unsubstituted or substituted

heterocyclic group or a lower alkyl group, or repre- sents hydroxy(lower) alkyl or carbamoyloxy(lower) alkyl,

R ₅ and R _g independently of one another represent hydrogen, hydroxy, hydroxy(lower) alkyl, cyano, amino, carbamoyl, carbamoyl (lower) alkyl, cyano(lower) alkyl, mono- or di-

(lower) alkylcarbamoyl, carbamoyloxy, ureido, amino (lower)alkyl, carbamoyloxy(lower)alkyl, mono- or di-

(lower)alkylcarbamoyl-(lower)alkyl, ureido(lower)alkyl, or a group of formula -CO-N or -CH ₂ C0-N J wherein

-N ) denotes a unsubstituted or substituted 3- to 6-

membered heterocyclic group which can contain addition¬ al hetero atoms, provided that R ₅ and R _g cannot be hydrogen at the same time,

R ₉ represents hydroxy(lower)alkyl or carbamoyloxy, and is an integer of 1 to 6, provided that when m is 1 and X is 0, R ₄ is other than unsubstituted amino(-NH ₂ ) •

A compound of formula (I) , as defined below, having the partial structure (A) above is a novel compound which was not disclosed in the prior art up to now. It is identified that the compound (I) exhibits a superior antibacterial activity against both of gram-positive bacteria such as Staphylococcus aureus and gram-negative bacteria such as Pseudomonas aeruginosa and further shows a good stability to DHP-I enzyme.

Accordingly, it is an object of the present invention to provide a novel 2-(2-substituted pyrrolidin-4-yl)thio- carbapenem derivative represented by the following general formula (I) :

and pharmaceutically acceptable salts thereof, in which

R-^ represents hydrogen or (lower)alkyl,

R ₂ represents hydrogen or anion, R ₃ represents hydrogen or (lower)alkanimidoyl,

X II Z represents -C-R ₄ or R _g ,

X represents O or NH,

R ₄ represents amino or- heterocyclic amine group, each of which can be unsubstituted or substituted with a group

of formula a unsubstituted or substituted heterocyclic group or a lower alkyl group, or repre¬ sents hydroxy(lower)alkyl or carbamoyloxy(lower)alkyl, Re and Rg independently of one another represent hydrogen, hydroxy, hydroxy(lower)alkyl, cyano, amino, carbamoyl, carbamoyl(lower)alkyl, cyano(lower)alkyl, mono- or di- (lower) alkylcarbamoyl, carbamoyloxy, ureido, amino (lower)alkyl, carbamoyloxy(lower)alkyl, mono- or di- (lower)alkylcarbamoyl-(lower)alkyl, ureido(lower)alkyl, or a group of formula -CO-N J or -CH ₂ CO-N J wherein -N J denotes a unsubstituted or substituted 3- to 6- membered heterocyclic group which can contain addition¬ al hetero atoms, provided that R ₅ and R _g cannot be hydrogen at the same time, R _g represents hydroxy(lower)alkyl or carbamoyloxy, and m is an integer of 1 to 6, provided that when m is 1 and X is O, R ₄ is other than unsubstituted amino(-NH ₂ ) •

Further, it is another object of the present invention to provide a process for preparation of the compound of formula(I) :

wherein R^ R ₂ , R ₃ , Z and m are defined as above, or salts thereof, which comprises reacting a compound of formula (II):

wherein R^ is as defined above and R ₇ represents a carboxy- protecting group, or a reactive derivative at the oxo group thereof or salts thereof with a compound of formula (III) :

wherein Z and m are as defined above and R _1Q represents an imino-protecting group, or salts thereof to obtain an in¬ termediate compound of formula (IV) ,

wherein R _{1 #} R ₇ , RI _Q . ^{~ anc} * ^~ * are as defined above, or salts thereof and subj ecting the resulting compound of formula ( IV) or salts thereof to elimination reaction of the car¬ boxy- and imino-protecting groups and, if necessary, react- ing the resulting compound of formula (I-b) ,

wherein R,, R ₂ , ^~ and m are as defined above, or salts thereof with a lower alkanimidoylating agent.

It is a further object of the present invention to provide a novel 2-substituted mercaptopyrrolidine compound represented by the following general formula (III) , which is an intermediate compound useful in the preparation of the desired compound of formula (I) :

wherein Z, R _1Q and m are as defined above, and a process for preparation thereof.

Further, it is another object of the present invention to provide a pharmaceutical composition containing a novel carbapenem compound of formula (I) as defined above.

BEST MODE FOR CARRYING OUT THE INVENTION

In one aspect, the present invention relates to a novel 2-(2-substituted pyrrolidin-4-yl)thio-carbapenem derivative of the following general formula (I) :

and pharmaceutically acceptable salts thereof, in which R~_ represents hydrogen or (lower) alkyl, R ₂ represents hydrogen or anion, R ₃ represents hydrogen or (lower) alkani idoyl,

X

II Z represents -C-R ₄ or.Rg,

X represents O or NH,

R ₄ represents amino or heterocyclic amine group, each of which can beunsubstituted or substituted with a group

of formula a unsubstituted or substituted heterocyclic group or a (lower) alkyl group, or repre¬ sents hydroxy(lower)alkyl or carbamoyloxy(lower)alkyl. Re and R _g independently of one another represent hydrogen, hydroxy, hydroxy(lower)alkyl, cyano, amino, carbamoyl, carbamoyl(lower)alkyl, cyano(lower)alkyl, mono- or di- (lower)alkylcarbamoyl, carbamoyloxy, ureido, amino- (lower) alkyl, carbamoyloxy(lower)alkyl, mono- or di- (lower)alkylcarbamoyl(lower)alkyl, ureido(lower)alkyl, or a group of formula -CO-N J or -CH ₂ CO-N wherein -N ) denotes a unsubstituted or substituted 3- to 6- membered heterocyclic group which can contain additional hetero atoms, provided that R ₅ and R _g cannot be hydro¬ gen at the same time, Rg represents hydroxy(lower)alkyl or carbamoyloxy, and m is an integer of 1 to 6, provided that when is 1 and X is 0, R ₄ is other than unsubstituted amino(-NH ₂ ) .

The compound of the present invention has the following basic structure :

This bas ic structure is named 7 -oxo-l-azabicyclo [ 3 . 2 . 0 ] hept-2-ene-2-carboxylic acid. In the present inven- tion , it is named l-carbapen-2-em-3-carboxylic acid accord¬ ing to the more generally and widely used nomenclature system for convenience.

The compound of formula (I) according to the present invention includes optical isomers due to asymmetric carbon atoms present in 1-, 5-, 6- and 8-position of the carbapenem moiety and side chain bound to 6-position thereof. Among such isomeric compound, the most preferred compound is a compound having trans configuration (5S, 6S) at 5 and 6 positions, R-configuration at 8-position and R-configuration at lower alkyl group (R ₁ ) present on 1-position, i.e. a compound having the whole configuration (1R, 5S, 6S, 8R) .

In addition, in a (2-substituted pyrrolidin-4-yl) thio group which is the side ^" chain present on 2-position of the carbapenem structure there may be isomeric compound due to asymmetric carbon atoms present on 2- and 4-positions of pyrrolidine, among which the compound having (2S',4S') or (2R',4R') configuration is most preferable. The compound of formula (I) according to the present invention also includes isomers at pyrrolidine moiety which is the side chain present on 2-position of carbapenem structure.

The desired compound of the present invention also includes a pharmaceutically acceptable salt of the compound of formula(I). Such pharmaceutically acceptable salt may include a base-addition salt such as an inorganic base salt, for example, an alkali metal salt (e.g. sodium salt, potas¬ sium salt, etc.), an alkaline earth metal salt (e.g. agne- sium salt, calcium salt, etc.), etc. or an organic base salt, for example, a salt with an organic base (e.g. triethylamine salt, dicyclohexylamine salt, ethanolamine salt, pyridine salt, picoline salt, etc.); an acid-addition salt such as an inorganic acid addition salt (e.g. hydro- chloride, hydrobromide, sulfate, phosphate, etc.) or an organic acid addition salt (e.g. formate, . acetate, tartrate, benzenesulfonate, etc.); an intermolecular quaternary salt, and the like.

The present invention also provides a process for preparation of 2- (2-substituted pyrrolidin-4-yl) thio-carba¬ penem derivatives of formula (I) as defined above and salts thereof. The process for preparation of the desired com-

pound of formula(I) according to the present invention can be illustrated by the following reaction scheme:

Process 1)

OD or a reactive derivative at the oxo group thereof or salts thereof

(TV) or salts thereof

Process 2.

ng

(I-a) or salts thereof

Process 3 )

Elimination of (CH ₂ ) _m — Z imino-protecting _â–  group(R ₁₀ )

(l-b) or salts thereof

Process 4)

(I-b) or salts thereof

^{) o} salts thereof

In the above reaction scheme,

R ₁ , R ₂ , R ₃ , Z and m are as defined above,

R ₇ represents a carboxy-protecting group and

R ₁₀ represents an imino-protecting group.

The term "lower alkyl" as used herein is intended to mean straight or branched alkyl having 1 to 6 carbon atoms and may include, for example, methyl, ethyl, propyl, isopro¬ pyl, butyl, s-butyl, t-butyl, pentyl, hexyl, etc. Prefera- ble lower alkyl is methyl, ethyl, propyl or t-butyl. Suit¬ able "carbamoyl (lower) alkyl" may include, for example, carbamoylmethyl, carbamoylethyl, carbamoylpropyl, l-(carbam- oylmethyl)ethyl, 1-carbamoyl-l-methylethyl, carbamoylbutyl, 1, l-dimethyl-2-carbamoylethyl, carbamoylpentyl, carbamoyl- hexyl, and the like, in which preferable one is carbamoyl- (C ₁ -C ₄ )alkyl and the most preferable one is carbamoylmeth¬ yl, carbamoylethyl or carbamoylpropyl. Suitable "lower alk- ani idoyl" may include straight or branched lower alkani id- oyl having 1 to 6 carbon atoms such as formimidoyl, aceti- midoyl, propionimidoyl, butyrimidoyl , isovalerimidoyl, pentanimidoyl, hexanimidoyl, and the like, in which the most preferable one is formimidoyl or acetimidoyl.

Suitable caboxy-pro ^' tecting group for R ₇ may include a group which can form esterified carboxy. Preferable exam¬ ples of the ester moiety of an esterified carboxy may in¬ clude lower alkyl ester such as methyl ester, ' ethyl ester, propyl ester, isopropyl ester, butyl ester, isobutyl ester, t-butyl ester, pentyl ester, hexyl ester, etc. ; lower alka- noyloxy(lower)alkyl ester which may have suitable substitu¬ ent(s) such as acetoxymethyl ester, propionyloxymethyl ester, butyryloxymethyl ester, valeryloxymethyl ester, pivaloyloxymethyl ester, hexanoyloxy ethyl ester, l(or 2)- acetoxyethyl ester, l(or 2, or 3)-acetoxypropyl ester, l(or 2, or 3 , or 4) -acetoxybutyl ester, l(or 2) -propionyloxy ethyl ester, l(or 2, or 3)-propionyloxypropyl ester, l(or 2)-butyryl oxyethyl ester, l(or 2)-isobutyryloxyethyl ester, l(or 2) -pivaloyloxy ethyl ester, l(or 2) -hexanoyloxyethyl

ester, isobutyryloxymethyl ester, (2-ethylbutyryl)oxymethyl ester, (3 , 3-dimethylbutyryl) oxymethyl ester, l(or 2)- pentanoyloxyethyl ester, etc.; (lower)alkanesulfonyl(lower) alkyl ester; mono-(or di- or tri-)halo(lower) alkyl ester such as 2-iodoethyl ester, 2,2,2-trichloroethyl ester, etc.; lower alkoxycarbonyloxy(lower)alkyl ester such as ethoxy- carbonyloxymethyl ester, 2-methoxycarbonyloxyethyl ester, 1- ethoxycarbonyloxyethyl ester, 1-isopropoxycarbonyloxyethyl ester, etc.; phthalidinyl(lower)alkyl ester; (5-lower alkyl- 2-oxo-l,3-dioxolan-4-yl) (lower)alkyl ester such as (5-meth- yl-2-oxo-l,3-dioxolan-4-yl) ethyl ester, (5-ethyl-2-oxo-l,3- dioxolan-4-yl)methyl ester, (5-propyl-2-oxo-l,3-dioxolan-4- yl) ethyl ester, etc.; lower alkenyl ester such as vinyl ester, allyl ester, etc. ; lower alkynyl ester such as ethy- nyl ester, propynyl ester, etc.; or aromatic(lower) alkyl ester which may have suitable substituent(s) , for example, benzyl ester, 4-methoxybenzyl ester, 4-nitrobenzyl ester, phenylethyl ester, trityl ester, benzhydryl ester, bis(methoxyphenyl)methyl ester, 3,4-dimethoxybenzyl ester, 4-hydroxy-3 ,5-t-butylbenzyl ester and the like; or aryl ester which may have suitable substituents, for example, phenyl ester, 4-chlorophenyl ester, tosyl ester, t-butyl- phenyl ester, mesityl ester, cumenyl ester, phthalidyl ester, and the like. Among these ester moieties, particu- larly preferable one may be substituted aromatic(lower)alkyl ester, most preferably 4-nitrobenzyl or phenyl(C.,-C ₄ )alkyl ester.

Suitable imino-protecting group for R, _Q may include acyl group such as aliphatic acyl substituted with aromatic or heterocyclic group derived from carboxylic acid, carbonic acid, sulfonic acid or carbamic acid, or carbamoyl, aliphat¬ ic acyl, aromatic acyl or heterocyclic acyl. Suitable aliphatic acyl may include saturated or unsaturated acyclic or cyclic acyl groups, for example, lower alkanoyl such as formyl, acetyl, propionyl, butyryl, isobutyryl, valeryl, isovaleryl, pivaloyl, hexanoyl, etc.; (lower)alkylsulfonyl such as mesyl, ethylsulfonyl, propylsulfonyl, isopropylsul-

fonyl, butylsulfonyl, isobutylsulfonyl, pentylsulfonyl, hexylsulfonyl, etc.; N-alkylcarbamoyl such as methylcarbam¬ oyl, ethylcarba oyl, etc.; (lower) alkoxycarbonyl such as methoxycarbonyl, ethoxycarbonyl, propoxycarbonyl, butoxycar- bonyl, t-butoxycarbonyl, etc.; (lower)alkenyloxycarbonyl such as vinyloxycarbonyl, allyloxycarbonyl, etc.; (lower) alkenoyl such acryloyl, methacryloyl, crotonoyl, etc.; cyclo(lower)alkylcarbonyl such as cyclopentylcarbonyl, ^' cyclo- propylcarbonyl, cyclohexylcarbonyl, etc., and the like. Suitable aromatic acyl may include heterocyclic carbonyl such as furoyl, thienylcarbonyl, nicotinoyl, isonicotinoyl, thiazolylcarbonyl, thiadiazolylcarbonyl, tetrazolylcarbonyl, etc. The aliphatic acyl substituted with aromatic group may include aralkanoyl such as phenyl(lower)alkanoyl, for exam- pie, phenylacetyl, phenylpropionyl, phenylhexanoyl, etc.; aralkoxycarbonyl such as phenyl(lower)alkoxycarbonyl, for example, benzyloxycarbonyl, phenylethyloxycarbonyl, etc.; aryloxyalkanoyl such as phenoxy(lower)alkanoyl, for example, phenoxyacetyl, phenoxypropionyl, etc., and the like. The aliphatic acyl substituted with heterocyclic group may include heterocyclic(lower)alkanoyl such as thienylacetyl, imidazolylacetyl, pyridylacetyl, tetrazolylacetyl, thiazoly- lacetyl, thiadiazolylacetyl, thienylpropionyl, thiadiazolyl- propionyl, tec. The above-mentioned acyl groups may have one or more substituent(s) and the preferable example of suitable substituent is as follows: (lower) alkyl such as methyl, ethyl, propyl, isopropyl, butyl, pentyl, hexyl, etc.; halogen such as chlorine, bromine, iodine, fluorine, etc.; (lower) alkoxy such as methoxy, ethoxy, propoxy, iso- propoxy, butoxy, pentyloxy, hexyloxy, etc.; (lower)alkylthio such as methylthio, ethylthio, propylthio, isopropylthio, butylthio, pentylthio, hexylthio, etc.; mono-(or di- or tri- )haloalkanoyl such as chloroacetyl, bromoacetyl, dichlo- roacetyl, trifluoroacetyl, etc.; mono-(or di- or tri-)haloal- koxycarbonyl such as chloromethoxycarbonyl, dichloromethoxy- carbonyl, 2,2,2-trichloro-ethoxycarbonyl, etc.; or nitro(or halo or lower alkoxy) aryl-oxycarbonyl such as nitrobenzy- loxycarbonyl, chlorobenzyloxycarbonyl, methoxybenzyloxycar-

bonyl, etc., and the like. More preferable example of imino-protecting groups as defined above may be (C ₂ ^_ C ₄ )alkenyloxycarbonyl or phenyl ( C^C^ )alkoxycarbonyl which may have nitro substituent, and the most preferable one may be allyloxycarbonyl or 4-nitrobenzyloxycarbonyl.

Suitable "lower alkylene" may include, for example, methylene, ethylene, trimethylene, tetramethylene, penta- methylene, hexamethylene, methylmethylene, ethylethylene, propylene, etc. , in which more preferable example may be C-, -

C ₄ alkylene and the most preferable one may be methylene.

In the definitions of R ₅ and R _g above, -N ) denotes a

3- to 6-membered heterocyclic group which may contain addi- tional hetero atoms, preferably nitrogen atom, such as substituted or unsubstituted aziridinyl, azetidinyl, pyrro- lidinyl, piperidinyl or piperazinyl, in which the preferable substituent may be carbamoyl, (lower)alkyl, hydroxy(lower) alkyl, cyano(lower) alkyl, amino(lower) alkyl, carbamoyloxy (lower)alkyl or amino.

"Substituted or unsubstituted heterocyclic amine" suitable for R ₄ preferably means a N-containing heterocyclic group, particularly aziridinyl, azetidinyl, pyrrolidinyl, piperidinyl or piperazinyl, which may be unsubstituted or substituted with carbamoyl, (lower) alkyl, hydroxy(lower) alkyl, cyano(lower)alkyl, amino(lower)alkyl, carbamoyloxy (lower)alkyl, ureido(lower)alkyl, carbamoyl(lower)alkyl, or mono or di-(lower)alkylcarbamoyl(lower)alkyl.

Among 2-(2-substituted pyrrolidin-4-yl)thio-carbapenem compounds of formula(I) according to the present invention, the preferable one is the compounds wherein R, represents hydrogen or lower alkyl, R ₂ represents hydrogen or anion, R ₃ represents hydrogen or straight or branched (C,-C ₈ )alkani-

midoyl, Z represents -

wherein R ₅ and Rg independently of one another represent hydrogen, hydroxy, hydroxy(lower) alkyl, CN, carbamoyl (lower) alkyl, cyano (lower) alkyl, carbamoyloxy(lower) alkyl, ureido (lower) alkyl or substituted or unsubstituted piperazinyl provided that R ₅ and Rg cannot be hydrogen at the same time, Rg represents hydroxy(C ₁ -C ₈ ) alkyl or carbamoyloxy and m is an integer of 1 to 4.

Particularly preferable compounds of formula(I) accord- ing to the present invention are the compounds wherein R, represents methyl, R ₂ represents hydrogen, R ₃ represents

0 II hydrogen or acetimidoyl, ^' Z represents -C-R ₄ or R _g , R ₄ repre-

sents , wherein R ₅ and R _g independently of one another represent hydrogen, hydroxy, hydroxy(C,-C ₄ ) alkyl, cyano, cyano (C ₁ -C ₄ ) alkyl , carbamoyloxy(C- _^ -C^ alkyl, ureido ( C^C^ ) alkyl, or piperazinyl optionally mono-substituted with substituent selected from carbamoyl, (C,-C ₄ )alkyl, hydroxy(C-_-C ₄ )alkyl, cyano(C- _^ -C^alkyl, amino(C- _^ -C^)alkyl, carbamoyloxy(C-^-C^alkyl, ureido(C ₁ ~C ₄ )alkyl, carbamoyl(C^ C ₄ )alkyl and mono- or di- (C^C^ )alkylcarbamoyl (C-^-C^ )alkyl, provided that R ₅ and Rg cannot be hydrogen at the same time, R _g represents hydroxy(C ₁ -C ₄ )alkyl or carbamoyloxy and m is an integer of 1 to 2.

The following example can be mentioned as the most preferable compound of formula(I) according to the present invention :

-(1R,5S, 6S) -2-[ (2S,4S) -2-{ (cyanomethylcarbamoyl)methylmerca- ptomethy1}pyrrolidin-4-y1]thio-6-[ (R)-1-hydroxyethy1]-l-me- thyl-l-carbapen-2-em-3-carboxylic acid, -(lR,5S,6S)-2-[ (2S,4S)-2-{ (cyanomethylcarbamoyl)ethylmercap- tomethyl}pyrrolidin-4-yl]thio-6-[ (R)-l-hydroxyethyl]-l-me- thyl-l-carbapen-2-em-3-carboxylie acid, -(1R,5S, 6S) -2-[ (2S,4S) -2-{ (aminoethylcarbamoyl)ethylmercap-

tomethyl}pyrrolidin-4-yl]thio-6-[ (R)-l-hydroxyethyl]-l-me- thyl-l-carbapen-2-em-3-carboxylic acid, -(1R,5S,6S)-2-[ (2S,4S) -2-{ (carbamoylmethylcarbamoyl)ethylme- rcaptomethyl}pyrrolidin-4-yl]thio-6-[ (R)-l-hydroxyethyl]-l- methyl-l-carbapen-2-em-3-carboxylic acid,

-(1R,5S,6S)-2-[ (2S,4S)-2-{ (hydroxyethylcarbamoyl)methylmer- captomethyl}pyrrolidin-4-yl]thio-6-[ (R)-1-hydroxyethyl]-l- methyl-l-carbapen-2-em-3-carboxylic acid, -(1R,5S,6S)-2-[ (2S,4S)-2-{ (carbamoylmethylcarbamoyl)methyl- mercaptomethyl}pyrrolidin-4-yl]thio-6-[ (R)-1-hydroxyethyl]- l-methyl-l-carbapen-2-em-3-carboxylic acid, -(1R,5S, 6S)-2-[ (2S,4S)-2-{ (cyanoethylcarbamoyl)ethylmercap- tomethyl}pyrrolidin-4-yl]thio-6-[ (R)-1-hydroxyethyl]-1-me- thyl-l-carbapen-2-em-3-carboxylie acid, -(1R,5S,6S)-2-[ (2S,4S)-2-{ (hydroxyethylcarbamoyl)ethylmerca- ptomethyl}pyrrolidin-4-yl]thio-6-[ (R)-1-hydroxyethyl]-1-me- thyl-l-carbapen-2-em-3-carboxylic acid, -(lR,5S,6S)-2-[ (2S,4S)-2-{ (carba oylethylcarbamoyl)ethylmer- captomethyl}pyrrolidin-4-yl]thio-6-[ (R)-l-hydroxyethyl]-l- methyl-l-carbapen-2-em-3-carboxylic acid,

-(1R,5S, 6S)-2-[ (2S,4S)-l-acetimidoyl-2-{ (2-carbamoylethylca- rbamoyl)methylmercaptomethyl}pyrrolidin-4-yl]thio-6-[ (R)-1- hydroxyethyl]-l-methyl-l-carbapen-2-em-3-carboxylic acid, -(lR,5S,6S)-2-[ (2S,4S)-l-acetimidoyl-2-{ (2-carbamoylmethyl- carbamoyl)ethylmercaptomethyl}pyrrolidin-4-yl]thio-6-[ (R)- l-hydroxyethyl]-l-methyl-l-carbapen-2-em-3-carboxylie acid, -(1R,5S,6S)-2-[ (2S,4S)-2-{ (1,2-dihydroxyethylcarbamoyl)meth- ylmercaptomethyl}pyrrolidin-4-yl]thio-6-[ (R)-1-hydroxyeth- yl]-l-methyl-l-carbapen-2-em-3-carboxylic acid, -(1R,5S, 6S)-2-[ (2S,4S)-2-{ (l-hydroxy-2-cyanoethylcarbamoyl)- methylmercaptomethyl}pyrrolidin-4-yl]thio-6-[ (R)-1-hydro- xyethyl]-l-methyl-l-carbapen-2-em-3-carboxylic acid, -(1R,5S,6S)-2-[ (2S,4S)-2-{ (l-hydroxy-2-aminoethylcarbamoyl)- methylmercaptomethyl}pyrrolidin-4-yl]thio-6-[ (R)-1-hydro- xyethyl]-l-methyl-l-carbapen-2-em-3-carboxylic acid,

-(1R,5S,6S)-2-[ (2S,4S)-2-{ (l-hydroxy-2-carbamoylethylcarba- moyl)methylmercaptomethyl}pyrrolidin-4-yl]thio-6-[ (R)-l- hydroxyethyl]-l-methyl-l-carbapen-2-em-3-carboxylic acid.

-(1R,5S,6S)-2-[ (2S,4S)-2-{ (1,2-dihydroxyethylcarbamoyl)eth- ylmercaptomethyl}pyrrolidin-4-yl]thio-6-[ (R)-1-hydroxyeth¬ yl]-l-methyl-l-carbapen-2-em-3-carboxylic acid,

-(1R,5S,6S)-2-[ (2S,4S)-2-{ (l-hydroxy-2-cyanoethylcarbamoyl)- ethylmercaptomethyl}pyrrolidin-4-yl]thio-6-[ (R)-1-hydroxy¬ ethyl]-l-methyl-l-carbapen-2-em-3-carboxylie acid,

-(1R,5S,6S)-2-[ (2S,4S)-2-{ (l-hydroxy-2-carbamoylethylcarba- moyl)ethylmercaptomethyl}pyrrolidin-4-yl]thio-6-[ (R)-1- hydroxyethyl]-l-methyl-l-carbapen-2-em-3-carboxylic acid, -(lR,5S,6S)-2-[ (2S,4S)-2-{ (1-(hydroxymethyl) -2-hydroxyethyl¬ carbamoyl)methylmercaptomethyl}pyrrσlidin-4-yl]thio-6-[ (R)- l-hydroxyethyl]-l-methyl-l-carbapen-2-em-3-carboxylic acid,

-(lR,5S,6S)-2-[ (2S,4S)-2-{ (1-(hydroxymethyl)-2-carbamoyleth- ylcarbamoyl)methylmercaptomethyl}pyrrolidin-4-yl]thio-6- [ (R)-l-hydroxyethyl]-l-methyl-l-carbapen-2-em-3-carboxylic acid,

-(lR,5S,6S)-2-[ (2S,4S)-2-{ (1-(hydroxymethyl)-2-carbamoyleth- ylcarbamoy1)ethylmercaptomethyl}pyrrolidin-4-y1]thio-6-[ (R) -l-hydroxyethyl]-l-methyl-l-carbapen-2-em-3-carboxylic acid,

-(1R,5S,6S)-2-[ (2S,4S)-2-{ (1-(carbamoyl ethyl)-2-ureidoeth- ylcarbamoyl)methylmercaptomethyl}pyrrolidin-4-yl]thio-6- [ (R) -l-hydroxyethyl]-l-methyl-l-carbapen-2-em-3-carboxylic acid, -(lR,5S,6S)-2-[ (2S,4S)-2-{ (1-(carbamoylmethyl)-2-cyanoethyl- carbamoyl)ethylmercaptomethyl}pyrrolidin-4-yl]thio-6-[ (R)- l-hydroxyethyl]-l-methyl-l-carbapen-2-em-3-carboxylic acid,

-(lR,5S,6S)-2-[ (2S,4S)-2-{ (1-(carbamoylmethyl)-2-aminoethyl- carbamoyl)ethylmercaptomethyl}pyrrolidin-4-yl]thio-6-[ (R)- l-hydroxyethyl]-l-methyl-l-carbapen-2-em-3-carboxylic acid,

-(1R,5S,6S) -2- [ (2S,4S)-2-{ (2-ureidoethylcarbamoyl)ethylmer¬ captomethyl}pyrrolidin-4-yl]thio-6-[ (R)-l-hydroxyethyl]-l- methyl-l-carbapen-2-em-3-carboxylie acid,

-(lR,5S,6S)-2-[(2S,4S)-2-{ (N-methylcarbamoyl)ethylmercapto- methyl}pyrrolidin-4-yl]thio-6-[ (R)-l-hydroxyethyl]-l-meth- yl-l-carbapen-2-em-3-carboxylie acid,

-(lR,5S,6S)-2-[ (2S,4S)-2-{ (N, -dimethylcarbamoyl)methylmer¬ captomethyl}pyrrolidin-4-yl]thio-6-[ (R)-l-hydroxyethyl]-l-

methyl-l-carbapen-2-em-3-carboxylie acid,

-(1R,5S,6S)-2-[ (2S,4S)-2-{l-acetimidoyl-2-(carbamoyl)ethyl- mercaptomethyl}pyrrolidin-4-yl]thio-6-[ (R)-1-hydroxyethy1]- 1-methyl-l-carbapen-2-em-3-carboxylie acid, -(lR,5S,6S)-2-[ (2S,4S) -2-{l-acetimidoyl-2-(N-methylcarbamo¬ yl)ethylmercaptomethyl}pyrrolidin-4-y1]thio-6-[-(R)-1-hydro- xyethyl]-l-methyl-l-carbapen-2-em-3-carboxylic acid,

-(lR,5S,6S)-2-[ (2S,4S)-2-{ (N-(2-hydroxyethyl) -piperazinylca- rbonyl)ethylmercaptomethyl}pyrrolidin-4-yl]thio-6-[ (R)-1- hydroxyethyl]-l-methyl-l-carbapen-2-em-3-carboxylic acid,

-(1R,5S,6S)-2-[ (2S,4S)-2-{ (N-(2-carbamoyloxyethyl)-piperazi- nylcarbonyl)ethylmercaptomethyl}pyrrolidin-4-yl]thio-6-[ (R) -l-hydroxyethyl]-l-methyl-l-carbapen-2-em-3-carboxylic acid, -(1R,5S,6S)-2-[ (2S,4S)-2-{ (N-(2-aminoethyl)-piperazinylcar- bonyl)ethylmercaptomethyl}pyrrolidin-4-yl]thio-6-[ (R)-1-hy- droxyethyl]-l-methyl-l-carbapen-2-em-3-carboxylic acid,

-(1R,5S,6S) -2-[ (2S,4S)-2-{ (2-hydroxyethyl)mercaptomethyl}py- rrolidin-4-yl]thio-6-[ (R)-1-hydroxyethyl]-1-methyl-l-carba- pen-2-em-3-carboxylic acid,

-(1R,5S,6S)-2-[ (2S,4S)-2-{ (3-hydroxypropyl)mercapto ethyl}- pyrrolidin-4-yl]thio-6-[ (R)-1-hydroxyethyl]-1-methyl-l-car- bapen-2-em-3-carboxylic acid,

-(1R,5S,6S)-2-[ (2S,4S)-2-{ (3-(carbamoyloxy)propyl)mercapto- methyl}pyrrolidin-4-yl]thio-6-[ (R)-1-hydroxyethyl]-1-meth- yl-l-carbapen-2-em-3-carboxylic acid,

-(1R,5S,6S)-2-[ (2S,4S)-2-=-{2-(hydroxyethylpiperidinylcarbony- lmethylcarbamoyl)ethylmercaptomethyl}pyrrolidin-4-yl]thio- 6-[ (R)-1-hydroxyethyl]-l-methyl-l-carbapen-2-em-3-carboxy- lie acid,

-(1R,5S,6S)-2-[ (2S,4S)-2-{2-(aminoethylpiperidinylcarbonyl- methylcarbamoyl)ethylmercaptomethyl}pyrrolidin-4-yl]thio- 6-[ (R)-l-hydroxyethyl]-l-methyl-l-carbapen-2-em-3-carboxy- lic acid, -(1R,5S,6S)-2-[ (2S,4S)-2-{2-(methoxyethylpiperidinylcarbon- ylmethylcarbamoyl)ethylmercaptomethyl}pyrrolidin-4-yl]thio- 6-[ (R)-l-hydroxyethyl]-l-methyl-l-carbapen-2-em-3-carboxy- lic acid.

-(1R,5S,6S)-2-[ (2S,4S)-2-{2-(carbamoyloxyethylpiperidinyl- methylcarbamoyl)ethylmercaptomethyl}pyrrolidin-4-yl]thio-6- [ (R)-l-hydroxyethyl]-l-methyl-l-carbapen-2-em-3-carboxylic acid, -(1R,5S,6S)-2-[ (2S,4S)-2-{2-(ureidoethylpiperidinylmethyl carbamoyl)ethylmercaptomethyl}pyrrolidin-4-yl]thio-6-[ (R)- l-hydroxyethyl]-l-methyl-l-carbapen-2-em-3-carboxylic acid, -(1R,5S,6S)-2-[ (2S,4S)-2-{2-(methoxymethylcarbamoyl)ethyl- mercaptomethyl}pyrrolidin-4-yl]thio-6-[ (R)-l-hydroxyethyl]- l-methyl-l-carbapen-2-em-3-carboxylic acid,

-(1R,5S,6S)-2-[ (2S,4S)-2-{2-(carbamoyloxymethylcarbamoyl)et¬ hylmercaptomethyl}pyrrolidin-4-yl]thio-6-[ (R)-l-hydroxyeth¬ yl]-l-methyl-l-carbapen-2-em-3-carboxylic acid, -(1R,5S,6S)-2-[ (2S,4S)-2-{2-(ureidomethylcarbamoyl)ethylmer- captomethyl}pyrrolidin-4-yl]thio-6-[ (R)-l-hydroxyethyl]-1- methyl-l-carbapen-2-em-3-carboxylic acid, -(1R,5S,6S)-2-[ (2S,4S)-2-{2-(methoxymethyloxymethylcarbamo¬ yl)ethylmercaptomethyl}pyrrolidin-4-yl]thio-6-[ (R)-l-hydro¬ xyethyl]-1-methyl-l-carbapen-2-em-3-carboxylie acid, -(lR,5S,6S)-2-[(2S,4S)-2-{2-(aminomethylcarbamoyl)ethylmer- captomethyl}pyrrolidin-4-yl]thio-6-[ (R)-l-hydroxyethyl]-1- methyl-l-carbapen-2-em-3-carboxylic acid, -(lR,5S,6S)-2-[ (2S,4S)-2-{2-(cyanoethylpiperidinylcarbonyl- methylcarbamoyl)ethylmercaptomethyl}pyrrolidin-4-yl]thio- 6-[ (R)-l-hydroxyethyl]-l-methyl-l-carbapen-2-em-3-carboxy- lic acid, -(1R,5S,6S)-2-[ (2S,4S)-2-{2-(methylcarbamoyloxymethylcarba- moyl)ethylmercaptomethyl}pyrrolidin-4-yl]thio-6-[ (R)-l-hy¬ droxyethyl]-l-methyl-l-carbapen-2-em-3-carboxylic acid, -(1R,5S,6S)-2-[ (2S,4S)-2-{2-(methoxymethyloxyethylcarbamo¬ yl)ethylmercaptomethyl}pyrrolidin-4-yl]thio-6-[ (R)-l-hy¬ droxyethyl]-l-methyl-l-carbapen-2-em-3-carboxylic acid, -(1R,5Ξ, 6S)-2-[ (2S,4S)-1-formimidoyl-2-{ (hydroxyethylcarbo¬ nyl)methylmercaptomethyl}pyrrolidin-4-yl]thio-6-[ (R)-1-hy- droxyethyl]-l-methyl-l-carbapen-2-em-3-carboxylic acid, and -(1R,5S,6S) -2-[ (2S,4S)-2-{ (carbamoylethyl) ercaptomethyl}py- rrolidin-4-yl]thio-6-[ (R)-l-hydroxyethyl]-1-methyl-l-carba- pen-2-em-3-carboxylic acid.

The novel desired- compound of formula(I) according to the present invention can be prepared by the processes as illustrated by Processes 1 to 4 above. The processes are explained in detail in the following.

(1) Process 1 :

In Process 1 of the method according to the present invention, a carbapenem derivative of formula(II) or a reac- tive derivative at the oxo group thereof or a salt thereof is reacted with a mercaptopyrrolidine derivative of formula(III) or a salt thereof to prepare a compound of formula(IV) or a salt thereof. In this reaction, the car¬ bapenem derivative of formula(II) can be preferably convert- ed into a reactive derivative of formula(Il-a) which is then reacted with the mercaptopyrrolidine derivative of formula(III) . Such reactive derivative can be prepared by reacting the compound of formula(II) with an acylating agent as illustrated in the following reaction scheme :

or salts thereof or salts thereof

In the above reaction scheme, R- _^ and R ₇ are as defined above, R _g represents acyl group as exemplified for the imino-protecting group R ₁₀ or 0, 0-substituted phosphono group derived from, for example, organic phosphoric acid as mentioned below.

Suitable acylating agent which can be used in the above

reaction may include conventional ones which can introduce the acyl group as mentioned above into the compound(II) . Preferable acylating agents may be organic sulfonic acid, organic phosphoric acid, or its reactive derivative such as acid halide or acid anhydride, for example, arenesulfonyl halide such as benzenesulfonyl chloride, p-toluenesulfonyl chloride, p-nitrobenzenesulfonyl chloride, p-bromobenzene- sulfonyl chloride, etc. ; arenesulfonic anhydride such as benzenesulfonic anhydride, p-toluenesulfonic anhydride, p- nitrobenzenesulfonic anhydride, etc.; optionally halogen- substituted(lower)alkanesulfonyl halide such as methanesul- fonyl chloride, ethanesulfonyl chloride, trifluoromethane sulfonyl chloride, etc. ; optionally halogen-substituted (lower) alkanesulfonic anhydride such as ethanesulfonic anhydride, ethanesulfonic anhydride, trifluoromethanesulfon¬ ic anhydride, etc.; di(lower)alkyl phosphorohaloridate such as diethyl phosphorochloridate, etc. ; diary1 phosphorohalor¬ idate such as diphenyl phosphorochloridate, etc., and the like, with diphenyl phosphorochloridate being most prefera- ble.

This acylation reaction for converting the compound of formula(II) into the reactive derivative of formula(Il-a) is preferably carried out in the presence of a solvent. For this purpose, any conventional organic solvent which does not adversely influence the reaction, for example, acetone, dioxane, acetonitrile, chloroform, dichloromethane, benzene, toluene, hexamethylphosphoramide, dichloroethane, tetrahy¬ drofuran, ethyl acetate, dimethylsulfoxide, N,N-dimethylfor- mamide, pyridine, etc. , can be used. Particularly, the most preferable solvent may be acetonitrile or benzene.

When the acylating agent is used in the form of a free acid or its salt in this acylation reaction, the reaction is usually carried out in the presence of a condensing agent. Suitable condensing agent for this purpose may include, for example, carbodiimide compounds such as N,N-diethylcarbodii- mide, N,N-diisopropylcarbodiimide, N,N-dicyclohexylcarbo-

dii ide, N-cyclohexyl-N'.-morpholinoethylcarbodiimide, N-cy- clohexyl-N'-(4-diethylaminocyclohexyl)carbodiimide, N-ethyl- N'-(3-dimethylaminopropyl)carbodii ide, etc.; imidazole com¬ pounds such as N,N'-carbonyldiimidazole, N,N'-carbonylbis(2- methylimidazole) , etc.; keteneimine compounds such as penta- methyleneketene-N-cyclohexylimine, diphenylketene-N-cyclohe- xylimine, etc.; l-alkoxy-1-chloroethylene; ethoxyacetylene; ethyl polyphosphate; isopropyl polyphosphate; phosphorus oxychloride; phosphorus trichloride; thienyl chloride; oxalyl chloride; a combination of triphenylphosphine with carbon tetrachloride or diazenedicarboxylate; 2-ethyl-7- hydroxybenzeneisoxazolium salt, and the like, with N,N'- carbonyldiimidazole being most preferable. In addition, this acylation reaction may be practiced in the presence of an inorganic or organic base. Suitable bases for this purpose may include hydroxides, carbonates, bicarbonates or alkanoates of an alkali metal such as lithium, sodium, potassium, etc. , and an alkaline earth metal such as calci¬ um, magnesium, etc. , for example, sodium hydroxide, potassi- u hydroxide, sodium carbonate, potassium carbonate, sodium bicarbonate, potassium bicarbonate, sodium acetate, magnesi¬ um carbonate or calcium carbonate; or tri(lower)alkyl amine such as triethylamine, trimethylamine, N,N-diisopropyl-N- ethylamine, etc.; pyridine compounds such as pyridine, picoline, lutidine, etc.; N,N-di(lower)alkylaminopyridine; N-(lower)alkylmorpholine; N,N-di(lower)alkylbenzeneamine, and the like. The most preferable base is N,N-diisopropyl- ethylamine or triethylamine.

The acylation reaction is generally carried out under cooling to warming, for example, at the temperature of -40°C to 50°C, preferably at the temperature of -20°C to 20°C. The reaction time is generally in the range of 0.5 to 3 hours, preferably in the range of 1 to 2 hours. In this acylation reaction, 1 to 3 moles, preferably 1 to 1.5 moles, of the base and 1 to 3 moles, preferably 1 to 1.5 moles of the acylating agent are generally used with respect to one mole of the compound of formula(II).

With regard to the compound(II) , it is well known that the 3,7-dioxo-l-azabicyclo[3.2.0]heptane ring system of the following formula(Il-b) lies in tautomeric relation with the 3-hydroxy-7-oxo-l-azabicyclo[3.2.0]hept-2-one ring system of the following formula(II-c) . Accordingly, it should be understood that both of these ring systems are substantially the same.

(II-b) (II-c)

The compound of formula (II) or the compound of formula(Il-a) or salts thereof can be subsequently reacted with the compound of formula(III) or salts thereof to pre¬ pare the intermediate compound of formula(IV) or salts thereof. In this reaction, the compound of formula(Il-a) produced by the above acylation reaction can be used with or preferably without isolation. The reaction of the compound (II) or (Il-a) or salts thereof with the compound(III) or salts thereof can be carried out in a reaction-inert solvent which does not adversely influence the reaction. As an example of such solvent, those solvent given in the explana¬ tion of the acylation reaction may be mentioned. The most preferable one may be acetonitrile or benzene. The reaction temperature can be varied within a substantially wide range. Generally, the reaction is carried out under cooling to warming.

(2) Process 2 :

In Process 2, the compound(IV) or salts thereof sub¬ jected to elimination reaction of the carboxy-protecting group R ₇ to prepare the compound(I-a) or salts thereof. The present reaction for removing the carboxy-protecting group is usually carried out by means of a conventional method such as hydrolysis, reduction, and the like.

(i) Hydrolysis

Hydrolysis for removing the carboxy-protecting group is preferably carried out in the presence of an acid or a base.

Suitable acid which can be used in such acid hydrolysis reaction may include an organic acid such as formic acid, acetic acid, propionic acid, trifluoroacetic acid, benzene- sulfonic acid, p-toluenesulfonic acid, etc. ; and an inorgan¬ ic acid such as hydrochloric acid, hydrobromic acid, sulfur¬ ic acid, phosphoric acid, etc. When the acidic hydrolysis is carried out using trifluoroacetic acid, the reaction can be accelerated by addition of a cation-trapping agent such as phenol, anisol, etc.

Suitable base for the basic hydrolysis may include an alkali metal hydroxide such as sodium hydroxide, potassium hydroxide, etc. ; an alkaline earth metal hydroxide such as magnesium hydroxide, calcium hydroxide, etc.; an alkaline earth metal hydride such as calcium hydride, etc. ; an alkali metal hydride such as sodium hydride, etc; an alkali metal alkoxide such as sodium methoxide, sodium ethoxide, potassi¬ um t-butoxide, etc.; an alkali metal carbonate such as sodium carbonate, potassium carbonate, etc.; or an alkali metal bicarbonate such as sodium bicarbonate, potassium bicarbonate, etc. , and the like.

This hydrolysis reaction is usually carried out in the presence of a solvent which does not adversely influence the reaction such as water, alcohol, for example, methanol, ethanol, etc., tetrahydrofuran, and the like, in which the preferable one may be methanol. When the acid or base used

in the hydrolysis is a liquid form, it can be also used as the solvent.

(ii) Reduction

The reduction method which can be used for elimination reaction of the carboxy-protecting group may include reduc¬ tion by using a combination of a metal such as zinc or zinc amalgam or a salt of chrome compound such as chromous chlo- ride or chromous acetate and an organic or inorganic acid such as acetic acid, propionic acid, hydrochloric acid, sulfuric acid, etc.; and conventional catalytic reduction in the presence of a conventional metallic catalyst such as palladium catalyst, for example, spongy palladium, palladium black, palladium oxide., palladium on carbon, palladium hydroxide on carbon, colloidal palladium, palladium on barium sulfate, palladium on barium carbonate, etc. , nickel catalysts, for example, reduced nickel, nickel oxide, Raney nickel, etc., or patinum catalyst, for example, platinum plate, platinum black, platinum on carbon, colloidal plati¬ num, platinum oxide, etc., and the like. Among those reduc¬ tion methods, the catalytic reduction using palladium oxide or palladium on carbon is most preferable. In case that the catalytic reduction is applied, the reaction is preferably carried out under neutral condition.

This reduction can be conveniently carried out in a solvent which does not adversely influence the reaction. For this purpose, suitable solvent may include, for example, water, alcohol such as methanol, ethanol, propanol, etc., dioxane, tetrahydrofuran, acetic acid, phosphate buffer, and the like, or a mixture thereof, with the mixed solvent of water and ethanol or water and tetrahydrofuran being most preferable. In this reaction, the preferable reaction temperature is in the range of 0°C to 30°C, most preferably in the range of 20°C to 30°C, and the reaction time is generally 0.5 to 4 hours and most preferably 2 to 3 hours. The amount of catalyst used in this reduction is usually

0.01 to 1 moles, preferably 0.01 to 0.5 moles, with respect to one mole of the compound of formula(IV) . Hydrogen atmos¬ phere is used in 1 to 4 atmospheric pressure and preferably in 3 to 4 atmospheric pressure.

In case that the carboxy-protecting group is allyl group, it can be deprotected by hydrogenolysis in the presence of a palladium compound. Suitable palladium com¬ pound used in this deprotecting reaction may include palla- dium on carbon, palladium hydroxide on carbon, palladium chloride, tetrakis(triphenylphospine)palladium (0), bis (dibenzylidenylacetone)palladium(O) , di[ (l,2-bis(diphenylph- osphino) ethane]palladium(0) , tetrakis(triphenylphosphite) alladium(O) , tetrakis(triethylphosphite)palladium(0) . This reaction can preferably be carried out in the presence of a scavenger of allyl group generated in situ. Suitable scav¬ enger which can be used for this purpose may include, for example, amine such as morpholine, N-methylaniline, etc.; an activated methylene compound such as dimedone, benzoylace- tate, 2-methyl-3-oxovaleric acid, etc.; a cyanohydrin com¬ pound such as o_-tetrahydropyranyloxybenzylcyanide, etc.; lower alkanoic acid such as formic acid, ammonium formate, sodium acetate, etc.; N-hydroxysuccinimide, and the like. This reaction can also be carried out in the presence of a base such as lower alkylamine, for example, butylamine, triethylamine, etc., pyridine, and the like. When palladi- um-ligand complex is used in this reaction, the reaction can preferably be carried out in the presence of the correspond¬ ing ligand, for example, triphenylphosphine, triphenylphos- phite, triethylphosphite, and the like. The reaction is usually carried out in a reaction-inert solvent such as water, methanol, ethanol, propanol, dioxane, tetrahydrofu¬ ran, acetonitrile, chloroform, dichloromethane, dichloroeth- ane, ethylacetate, etc. The reaction temperature can be varied within a substantially wide range and the reaction is generally carried out under cooling to warming.

The elimination reaction of protecting group can be

carried out using a reducing agent appropriately selected depending on the kind of carboxy-protecting group to be eliminated.

(3) Process 3 :

In the reaction of Process 3, the compound of formula(I-a) or salts thereof is subjected to elimination reaction of the imino-protecting group(R _1Q ) to prepare the compound of formula(I-b) ^' or salts thereof. This elimination reaction of the imino-protecting group can generally be carried out by means of a conventional method such as hydrol¬ ysis, reduction, and the like. The method of hydrolysis and reduction, and the reaction conditions (reaction tempera- ture, solvent, and the like) are substantially the same as those illustrated for elimination reaction of the carboxy- protecting group in Process 2 above. If necessary, the reaction of Process 3 can be directly carried out in the same reaction vessel without isolation of the compound(I-a) produced by Process 2.

The reaction of Process 3 for removing the imino- protection group is carried out in the same manner as the reaction of Process 2 for removing the carboxy-protecting group.

Alternatively, Process 3 can be practiced simultaneous¬ ly with Process 2 and this case is also included within the scope of the present invention.

(4) Process 4 :

In Process 4, the compound of formula (I-b) or salts thereof is reacted with lower alkanimidoylating agent to prepare the compound of formula (I) wherein R-. is lower alkanimidoyl group, i.e. the compound of formula(I-c) , or salts thereof.

In this reaction, suitable lower alkanimidoylating agent may be any of conventional ones which can introduce the lower alkanimidoyl group into the compound of formula (I-b) . As a specific example of such lower alkanimidoylat- ing agent, the following compounds can be mentioned : lower alkyl(lower)alkanimidate such as methyl formimidate, ethyl formimidate, methyl acetimidate, ethyl acetimidate, ethyl propionimidate, methyl butyrimidate, ethyl isovalerimidate, ethyl pentanimidate, ethyl hexanimidate, etc.; lower alkani- midoyl halide such as formimidoyl chloride, formimidoyl bromide, acetimidoyl chloride, aceti idoyl bromide, propion- imidoyl bromide, butyri idoyl chloride, isovalerimidoyl chloride, pentani idoyl chloride, hexanimidoyl chloride, etc. , and the like, with methyl formimidate or methyl ace- timidate being most preferable.

This reaction is usually carried out in a solvent which does not adversely influence the reaction such as tetrahy¬ drofuran, dioxane, water, methanol, ethanol, etc., or a mixture thereof. The most preferable solvent may be a mixed solvent of water and ethanol. The reaction temperature can be varied within a substantially wide range and the reaction is generally carried out under cooling to warming, most preferably at the temperature of -10°C to 10°C, for the reaction time of 0.5 to 3 hours, more preferably 1 to 1.5 hours.

In the present reaction, the lower alkanimidoylating agent may be used in an amount of 1 to 2 moles, preferably 1 to 1.5 moles and particularly 1.2 moles, with respect to one mole, of the compound(I-b) .

The reaction can also be carried out in the presence of an inorganic or organic base. Suitable base which can be used in this reaction may be those given in the explanation of the reaction of Process 1. In this reaction, the base is used in an amount sufficient to maintain the pH value of the reaction solution at the weak alkaline level, preferably in

the range of pH 8 to 9 and most preferably in the range o pH 8.5 to 8.7.

The compound of formula(I) and salts thereof obtaine from the Processes 1 to 4 according to the present inventio can be isolated and purified by means of a conventiona manner, for example, extraction, precipitation, fractiona crystallization, recrystallization, column chromatography, and the like.

Suitable salts of the compounds of formulae(II) , (II- a) , (III) , (IV) , (I-a) , (I-b) and (I-c) , which are used as starting materials and reactants or obtained as reaction products in the Processes 1 to 4 above, may be the same as those specifically exemplified in connection with the salts of the compound of formula(I).

The mercaptopyrrolidine derivative of formula(III) , which is used as a reactant in Process 1 above for prepara- tion of the compound(I) according to the present invention, is a novel compound which was never disclosed in the prior art. Accordingly, another object of the present invention is to provide a compound of the following formula(III) :

(CH ₂ ) _m —Z (III)

or salts thereof, in which R _1Q represents an imino-protecting group,

X II Z represents -C-R ₄ or Rg,

X represents O or NH,

R ₄ represents amino or heterocyclic amine group, each of which can be unsubstituted or substituted with a group

of formula , a unsubstituted or substituted heterocyclic group or a (lower)alkyl group, or repre¬ sents hydroxy(lower)alkyl or carbamoyloxy(lower)alkyl,

R ₅ and Rg independently of one another represent hydrogen, hydroxy, hydroxy(lower)alkyl, cyano, amino, carbamoyl, carbamoyl(lower)alkyl, cyano(lower)alkyl, mono- or di- (lower)alkylcarbamoyl, carbamoyloxy, ureido, amino(lo¬ wer) alkyl, carbamoyloxy(lower)alkyl, mono- or di-(lo- wer)alkylcarbamoyl(lower)alkyl, ureido(lower)alkyl, or a group of formula -CO-N J or -CH ₂ C0-N J wherein -N J denotes a unsubstituted or substituted 3- to 6- membered heterocyclic group which can contain addition¬ al hetero atoms, provided that R ₅ and R _g cannot be hydrogen at the same time,

R _g represents hydroxy(lower)alkyl or carbamoyloxy, and m is an integer of 1 to 6, provided that when m is 1 and X is 0, R ₄ is other than unsubstituted amino(-NH ₂ ) .

Further, the present invention provides a process for preparation of the compound of formula(III) and salts there¬ of. The process for preparation of the compound(III) ac¬ cording to the present invention can be represented by Methods A and B as depicted below.

Method A

(V) or a reactive derivative at the hydroxy group thereof or salts thereof

or salts thereof (ffl-a)

Method B

Elimination of CH ₂ - S-(CH ₂ ) _m -z ^mer capto-protecting

R„-S ^â–  Cc -Rio group(R ₁₁ )

or salts thereof (IH-a)

or salts thereof (III)

In the above reaction scheme, R _1;L represents a mercap- to-protecting group, and Rι _0/ ^~ and m are as defined above.

Suitable "mercapto-protecting group" for R _1;L may in-

elude acyl group as mentioned above in connection with the imino-protecting group; ar(lower)alkyl group such as ono- or di- or tri-phenyl (lower) alkyl, for example, benzyl, phenathyl, benzhydryl, trityl, etc.; and the like, in which more preferable example may be C ₁ ~C ₄ alkanoyl, aroyl and triphenyl(C ₁ -C ₄ ) alkyl and the most preferable mercapto- protecting group may be acetyl.

Methods A and B for preparing the novel compound of formula(III) and salts thereof are explained in detail in the following.

a) Method A :

In Method A, the compound of formula(V) or a reactive derivative at the hydroxy group thereof or salts thereof can be reacted with the mercaptan derivative of formula(VI) or salts thereof to prepare the derivative of formula(Ill-a) or salts thereof.

Suitable reactive derivative at the hydroxy group of the compound of formula(V) may include a conventional one such as halides, for example, chloride, bromide, iodide, etc. ; sulfonates, for example, methanesulfonate, benzenesul- fonate, toluenesulfonate, etc. ; and the like, with a sulfo¬ nate compound, particularly methanesulfonate being most preferable. In case that the compound of formula(V) is a methanesulfonate derivative, the reaction of Method A can be carried out, for example, by reacting one equivalent weight of the compound of formula (VI) with 1 to 2 equivalent weight, preferably' 1.2 equivalent weight, of the methane¬ sulfonate compound of formula (V) and 1 to 2 equivalent weight, preferably 1.2 equivalent weight, of an organic or inorganic base in a halogenated alkane solvent such as dichloromethane, at the temperature of -10°C to 40°C, more preferably -5°C to 0°C, for 1 to 3 hours, more preferably 1 to 1.5 hours.

Preferable example of the mercaptan derivative of formula (VI) used as a reactant in the reaction of Method A may be ar (lower) alkanethiol such as mono- or di- or tri- phenyl (lower) alkanethiol, for example, phenylmethanethiol, diphenylmethanethiol, triphenylmethanethiol, etc.; thio (lo¬ wer) alkanoic-S-acid such as thioacetic-S-acid, etc.; thioar- enoic-S-acid such as thiobenzoic-S-acid, etc.; and the like, in which more preferable example may be triphen ^' yl (C-_- C ₄ ) alkanethiol, thio (C-^-C^) alkanoic-S-acid and thio(Cg-C ₁₀ ) - arenoic-S-acid, and salts thereof. The most preferable mercaptan derivative (VI) may be thioacetic-S-acid or its potassium salt. In case that the compound (VI) is thioacet¬ ic-S-acid potassium salt, the reaction of Method A can be carried out in a stoichiometric manner, for example, by reacting one equivalent weight of the compound (V) with 1 to 2 equivalent weight, preferably 1.5 equivalent weight, of potassium thioacetate(VI) in a solvent such as dimethylsul- f oxide, hexamethylphosphoramide or N,N-dimethylf ormamide, preferably in N, N-dimethy If ormamide, at the temperature in the range of 60°C to 100°C, more preferably 80°C to 90°C, for 2 to 5 hours, more preferably 2.5 to 3 hours.

In Method A, when- the compound of formula (VI) is ar (lower) alkanethiol, the compound of formula (V) is prefera- bly used in the form of its reactive derivative at the hydroxy group and the reaction can be conveniently carried out in the presence of an inorganic or organic base. Suit¬ able inorganic or organic base used in this reaction may be those exemplified in Process 1 above.

In case that the compound (VI) is thio (lower) alkanoic-S- acid or thioarenoic-S-acid, the reaction of Method A is preferably carried out in the presence of a conventional condensing agent such as a combination of triarylphosphine, for example, triphenylphosphine, etc., and di ( lower) alkyla- zodicarboxylate, for example, diethylazodicarboxylate, etc. This reaction is usually carried out in a conventional solvent which does not 'adversely influence the reaction,

such as dichloromethane, methanol, ethanol, propanol, pyri¬ dine, N,N-dimethyIformamide or tetrahydrofuran, with tet¬ rahydrofuran being most preferable. In such a case, the reaction of Method A is preferably carried out, for example, by reacting one equivalent weight of the compound of formu¬ la(V) with 1 to 2 equivalent weight, preferably 1.5 equiva¬ lent weight, of thio(lower)alkanoic-S-acid such as thioacet- ic acid, 1 to 2 equivalent weight, preferably 1.5 equivalent weight, of diethylazodicarboxylate and 1 to 5 equivalent weight, preferably 2 equivalent weight, of triphenylphos¬ phine in tetrahydrofuran solvent at the temperature of -40°C to 10°C, preferably 0°C to 5°C, for 2 to 5 hours, preferably 2 to 3 hours.

As a result of the reaction of Method A, the configura¬ tion on the carbon atom substituted with the hydroxy group of the compound(V) is inverted in the resulting compound (Ill-a) .

* ^~ ) Method B :

In Method B, the compound of formula(Ill-a) or salts thereof obtained from Method A is subjected to elimination reaction of the mercapto-protecting group to prepare the compound of formula(III) or salts thereof.

The elimination reaction of the mercapto-protecting group according to Method B is carried out by means of a conventional method as described below and can be appropri- ately selected depending on the kind of the mercapto-pro¬ tecting group to be eliminated. For example, when the protecting group is ar(lower)alkyl group, this protecting group can generally be eliminated by treating with a silver compound such as silver nitrate, silver carbonate, etc. This elimination reaction with silver compound is preferably carried out in the presence of an organic base such as pyridine, etc. The resulting silver salt of the compound of formula(III) can be converted into its alkali metal salt, if

necessary, by treating with alkali metal halide such as sodium iodide, potassium iodide, etc.

Further, in case that the mercapto-protecting group is acyl,. this protecting group can generally be eliminated by solvolysis such as alcoholysis or hydrolysis using acid or base. Suitable acid and base which can be used in this reaction may be those exemplified in connection with hydrol¬ ysis in Process 2 above, in which sodium methoxide and sodium hydroxide is most preferable. This hydrolysis is usually carried out in a conventional solvent which does not adversely influence the reaction such as alcohol, for exam¬ ple, methanol, ethanol, etc., water, pyridine, N,N-dimethyl- formamide, etc., or a mixture thereof, in which the most preferable one is methanol or water.

In case that the reaction of Method B is carried out by basic hydrolysis, the reaction is carried out, for example, by using 1 to 2 moles, preferably 1 to 1.5 moles, of a base with respect to one mole of the compound of formula(Ill-a) in the presence of a solvent such as water, alcohol or tetrahydrofuran, most preferably in methanol solvent, at the temperature of -20°C to 50°C, most preferably -10°C to 10°C, for 0.5 to 2 hours, most preferably 0.5 to 1 hour. The reaction is practiced substantially in a stoichiometric manner.

The compound of formula(III) obtained from Methods A and B above can be isolated and purified according to a conventional method, for example, by extraction, precipita¬ tion, fractional crystallization, recrystallization, chroma¬ tography, and the like.

The novel carbapenem derivative of formula (I) and pharmaceutically acceptable salts thereof according to the present invention exhibit a potent antibacterial activity against various gram-positive and gram-negative bacterial strains and, therefore, has a clinical utility for prophy-

laxis and treatment of various bacterial infectious dis¬ eases. Antibacterial activity of the desired compoun (I) of the present invention is demonstrated by the following experiments including in vitro antibacterial activity test.

Test Examples

Test 1 : In vitro antibacterial activity test

The antibacterial activity of the compound(I) of the present invention was determined by measuring the minimal inhibitory concentration(MIC) according to the antibacterial activity test method as described below. The compounds of Examples 1,2,3 and 4 according to the present invention were used as a test compound and imipenem was used as a compara¬ tive compound.

The test compounds and comparative compound were gradu¬ ally diluted in Mueller Hinton Agar(MHA) medium. Test strains as listed in the following table were inoculated into the medium in an amount of 10° viable cells/ml and then incubated at 37°C for 18 hours. Then, MIC value was meas- ured and expressed in terms of . g/ml. The results are shown in the following Table 1.

Table 1. Minimal Inhibitory Concentration of the compound(I) and imipenem

From the result given in Table 1 above, it can be seen that the compound of formula(I) and pharmaceutically accept¬ able salts thereof according to the present invention exhib¬ it a considerably superior antibacterial activity in com-

parison with the known compound in the prior art, imipenem.

Test 2 : Stability test against kidney dehydrogenase DHP-I

Each of the novel compounds of Examples 1,2,3 and 4 of the present invention and the comparative compound imipenem was dissolved in lOmM phosphate buffer and the resulting solution was mixed with dehydrogenase enzyme solution ex- tracted from pig kidney in a ratio of equivalent amounts. The resulting mixture was incubated at 37°C for 2 hours, during which the decomposition degree of the compound was measured. The result thereof was given in the following Table 2 in terms of percentage of the remaining amount on the basis of the initially added amount of the compound.

Table 2. Stability of the compounds(I) and imipenem on DHP-I enzyme

(Unit:%)

Time(minute)

10 20 40 80 120

Test Compound

Imipenem 99.2 97.9 94.3 87.7 77.0 66.6

Compound of Example 1 99.7 99.4 99.1 98.1 97.5 97.2

Compound of Example 2 100 99.4 98.6 97.2 96.3 96.3

Compound of Example 3 99.7 99.2 98.9 98.0 97.2 97.0

Compound of Example 4 100 99.8 99.0 98.8 98.8 98.8

From the result given in Table 2 above, it can be seen that the known compound in prior art, imipenem, was decom-

posed by about 40% within about 2 hours from the begining of mixing incubation, while the novel compound of formula(I) of the present invention was substantially never decomposed and shows a high stability. Such test result demon trates that imipenem has a disadvantage in that it should be adminis¬ tered together with an enzyme inhibitor such as Cilastatin, whereas the compound of the present invention shows stably a high antibacterial activity even when it is administered alone.

As can be demonstrated by the results of Tests 1 and 2 above, the carbapenem derivative of formula(I) and pharma¬ ceutically acceptable salts thereof according to the present invention are clinically used for the purpose of prophylaxis and treatment of infectious diseases. For this purpose, the compound of formula(I) and pharmaceutically acceptable salts thereof can be formulated into a pharmaceutically acceptable preparation suitable for administration, together with pharmaceutically acceptable carriers such as solid or liquid excipients. Such pharmaceutical preparation may be in solid or liquid form, for example, tablet, capsule, granule, powder, pill, solution, suspension, syrup, emulsion, and the like. If necessary, the pharmaceutical preparation can additionally contain conventional additives, for example, formulating aid, stabilizer, wetting agent, thickening agent, disintegrator, perfume, pigment, binder, and the like.

The dosage of the compound of formula(I) and pharmaceu- tically acceptable salts thereof according to the present invention may vary depending on kind and severity of dis¬ eases to be treated, age and conditions of the patient, and kind of the compound(I) to be administered. In general, an amount of 100 to 500mg per day may be administered per one adult patient as a single dose or a multiple-divided dose.

The present invention will be explained by the follow¬ ing Examples and Reference Examples in more detail. Howev-

er, it should be understood that the present invention is not limited by these examples in any manner.

Example 1

Preparation of ( 1R, 5S , 6S) -2- [ (2S , 4S) -2-{ (cyanomethylcarbam¬ oyl ) methy lmercaptomethyl } pyrrolidin-4-yl ] thio-6- [ (R) -1- hydroxyethyl] -l-methyl-l-carbapen-2-em-3-carboxylic acid

(1) 580mg of 4-nitrobenzyl-2-diphenoxyphosphoryloxy-6- [ (R)-l-hydroxyethyl]-l-methyl-l-carbapen-2-em-3-carboxylate and 380mg of (2S,4S)-1-(p-nitrobenzyloxycarbonyl)-2-{ (cyano- methylcarbamoyl)methylmercaptomethyl}-4-mercaptopyrrolidine were dissolved in 25ml of acetonitrile. To the resulting solution was added dropwise 0.25ml of N,N-diisopropylethyla- mine under nitrogen atmosphere at -10°C to -5°C and then the reaction mixture was stirred overnight at 5°C. The reaction solution was concentrated under reduced pressure. 20ml of 5% sodium bicarbonate solution was added to the residue and the mixture was extracted with 30ml of ethylacetate. The extract was dried over magnesium sulfate and concentrated under reduced pressure. The residue was subjected to column chromatography(eluent: EtOAc:acetone=3 :2 v/v) to obtain 760mg of 4-nitrobenzyl (1R,5S,6S)-2-[ (2S,4S)-1-(p-nitroben¬ zyloxycarbonyl)-2-{ (cyanomethylcarbamoyl)methylmercaptometh¬ yl}pyrrolidin-4-yl]thio-6-[ (R)-l-hydroxyethyl]-1-methy1-1- carbapen-2-em-3-carboxylate.

IR(Nujol)cm~ ¹ : 1755, 1751, 1710, 1653

NMR(CDCl ₃ ) : 1.27(3H,d,J=8Hz) , 1.37 (3H,d,J=8Hz) , 1.78-

1.90(lH,m), 2.30-2.80(7H,m) , 3.31-3.76(4H,m) , 5.04-5.54(6H,m) , 7.44-7.50(2H,dd,J=18Hz) , 7.55-7.65(4H,d,J=9Hz) , 8.17-8.25(6H,m)

(2) 690mg of the compound obtained in the above (1) was dissolved in 10ml of tetrahydrofuran. To the resulting

solution were added 18ml. of 0.1M 4-morpholinopropane sulfo¬ nate solution (pH=7.0) , 1.8ml of ethanol and 0.3g of 10% palladium on carbon and then the mixture was hydrogenated under pressure. After the reaction time of 4 hours, the catalyst was filtered off and the filtrate was washed with 20ml of dichloromethane. The aqueous layer was separated and purified with polymer chromatography(CHP-20, eluent: 5%THF-H ₂ 0) . Then, the eluate was lyophilized to Obtain 170mg of the title compound.

Melting Point : 173-174.5 ^α C (dec.) IR(KBr)cm ^"1 : 1750, 1580, 1390

NMR(D ₂ 0) ξ : 1.20 (3H,d,J=8Hz) , 1.28 (3H,d,J=8Hz) , 1.40-2.10(2H,m) , 2.55-3.95(6H,m)

Example 2

Preparation of (IR, 5S, 6S) -2-[ (2S, 4S) -2-{ (cyanomethylcarbam- oyl) ethylmercaptomethyl}pyrrolidin-4-yl] thio-6- [ (R) -1- hydroxyethyl]-l-methyl-l-carbapen-2-em-3-carboxylic acid

(1) The reaction was carried out in the same manner as that of Example 1-(1) , starting from 480mg of 4-nitrobenzyl- 2-diphenoxyphosphoryloxy-6-[ (R) -l-hydroxyethyl]-1-methy1-1- carbapen-2-em-3-carboxylate and 320mg of (2S , 4S) -[ 1- (p- nitrobenzylojxycarbonyl) -2-{ (cyanomethylcarbamoyl) ethylmer- captomethyl}-4-mercapto]pyrrolidine to obtain 580mg of 4- nitrobenzyl (1R,5S, 6S)-2-[ (2S,4S) -1-(p-nitrobenzyloxycarbon- yl) -2-{ (cyanomethylcarbamoyl)ethylmercaptomethyl}pyrrolidin- 4-yl]thio-6-[ (R) -l-hydroxyethyl]-l-methyl-l-carbapen-2-em-3- carboxylate.

IR(Nujol)cm ^"1 : 1780, 1750, 1620

NMR(D ₂ 0) ξ : 1.21 (3H,d,J=8Hz) , 1.30 (3H,d,J=8Hz) , 3.02-3.85(12H,m) , 4.80(2H,br. s) , 5.25(2H,Ξ) , 5.38(2H,dd,J=18Hz) ,

7.58(2H,d,J=8Hz) , 8.26(6H,m)

(2) The compound obtained in the above (1) was hydrog¬ enated in the same manner as that of Example l-(2) to obtain 120mg of the title compound.

Melting Point : 175-175.5°C (dec.) IRfKBrJc ^"1 : 1750, 1580

NMR(D ₂ 0) £ : 1.23 (3H,d,J=7.9Hz) , 1.30 (3H,d,J=6.8Hz) , 1.81(lH,dd,J=18Hz,J=8.5Hz) , 2.70-3.34 (5H,m) ,

3.45-3.85(3H,m) , 4.25-4.27 (2H,m)

Example 3

Preparation of (IR, 5S, 6S) -2-[ (2S,4S) -2-{ (hydroxyethylcarbam¬ oyl) ethylmercaptomethyl} yrrolidin-4-yl ]thio-6-[(R)-i- hydroxyethyl]-l-methyl-l-carbapen-2-em-3-carboxylic acid

(1) The reaction was carried out in the same manner as that of Example 1-(1), starting from 550mg of 4-nitrobenzyl- 2-diphenoxyphosphoryloxy-6-[ (R) -l-hydroxyethyl] -1-methyl-l- carbapen-2-em-3-carboxylate and 410mg of (2S, 4S) -[ 1- (p- nitrobenzyloxycarbonyl) -2-{ (hydroxyethylcarbamoyl) ethylmer- captomethyl}-4-mercapto]pyrrolidine to obtain 420mg of 4- nitrobenzyl (1R,5S, 6S) -2-[ (2S,4S) -1- (p-nitrobenzyloxycarbon- yl) -2-{ (hydroxyethylcarbamoyl) ethylmercaptomethyl}pyrroli- din-4-yl ] thio-6- [ (R) -l-hydroxyethyl ] -l-methyl-l-carbapen-2- em-3-carboxylate.

IR(Nujol)cm ^"1 : 3400, 1770-1740, 1710-1680, 1605 NMR(CDC1 ₃ ) S = 1.1-1.6 (6H,m) , 5.1-5.6 (4H,m) , 7.3-7.7

(4H,m) , 8.21(4H,d,J=9Hz)

(2) The compound obtained in the above (1) was hydrog¬ enated in the same manner as that of Example l-(2) to obtain 250mg of the title compound.

Melting Point : 172-173°C (dec.) IR(KBr)cm ^_1 : 1765-1725, 1590-1550

NMR(D ₂ 0) 8 : 1.21 (3H,d,J=8Hz) , 1.28 (3H,d,J=7Hz) , 1.52-2.0(4H,m) , 2.49-2.9 (3H,m)

Example 4

Preparation of (IR, 5S, 6S) -2-[ (2S, 4S) -2-{ (2 -carbamoylmethyl¬ carbamoyl) ethylmercaptomethyl}pyrrolidin-4-yl] thio-6-[ (R) -1- hydroxyethyl]-l-methyl-l-carbapen-2-em-3-carboxylic acid

(1) The reaction was carried out in the same manner as that of Example 1-(1), starting from 650mg of 4-nitrobenzyl- 2-diphenoxyphosphoryloxy-6-[ (R) -l-hydroxyethyl ]-l-methyl-l- carbapen-2-em-3-carboxylate and 350mg of (2S , 4S) - [ 1- (p- nitrobenzyloxycarbonyl) -2-{ (2-carbamoylmethy lcarbamoyl) eth- ylmercapto_nethyl}-4-mercapto] pyrrolidine to obtain 450mg of 4 -nitrobenzyl (lR,5S,6S)-2-[ (2S,4S) -1- (p-nitrobenzy loxy car¬ bonyl) -2-{ (2-carbamoylmethylcarbamoyl) ethy lmercaptomethyl} pyrrolidin-4-yl ] thio-6- [ (R) -l-hydroxyethyl ] -1-methyl-l-car- bapen-2-em-3-carboxylate.

IR(Nujol) cm -1 1770, 1705, 1610, 1525

NMR(CDC1 ₃ ) £ : 1.25 (3H,d,J=7.5Hz) , 1.32 (3H,d,J=6Hz) ,

3.10-4.83 (3H,m) . 4.81(2m,br. s) , 5.24(2H,S), 5.38(2H,dd,J=18Hz) , 7.56-7.68(4H,dd,J=18Hz) , 8.24 (4H,d,J=8Hz)

(2) The compound obtained in the above (1) was hydrog¬ enated in the same manner as that of Example l-(2) to obtain 120mg of the title compound.

Melting Point : 168-172°C (dec.)

IR(KBr)cm~ ¹ : 1755, 1680

NMR(D ₂ 0) $ : 1.21 (3H, d, J=9Hz) , 1.27 (3H, d, J=6Hz) ,

1.42-2.03 (2H,m) , 2.53-4.36 (10H,m)

Example 5

Preparation of (IR, 5S, 6S) -2-[ (2S,4S) -l-acetimidoyl-2-{ (2-ca- rbamoyl ethy lcarbamoyl) ethy lmercaptomethyl} pyrrol idin-4-yl] thio-6- [ (R) -l-hydroxyethyl ] -l-methyl-l-carbapen-2-em-3-car- boxylic acid

120mg of the compound obtained in Example 4-(2) , (1R,5S, 6S) -2-[ (2S,4S) -2-{ ( 2 -carbamoylmethylcarbamoyl) ethy lm- ercaptomethyl}pyrrolidin-4-yl]thio-6-{ (R) -l-hydroxyethyl ]-l- methyl-l-carbapen-2-em-3.-carboxylic acid was dissolved in 30ml of distilled water. To the resulting solution was added 1.4g of ethylacetimidate hydrochloride. Then, the reaction mixture was adjusted to pH 8.4-8.6 by adding 10% aqueous sodium carbonate solution at 0 to 4°C. When the reaction was completed, the reaction mixture was adjusted to pH 6.5 by adding lN-aqueous hydrochloric acid solution and then washed with the mixture of 50ml of ethyl acetate and 10ml of tetrahydrofuran. The organic layer was removed and the aqueous layer was subjected to polymer chromatography (CHP-20, eluent : 5% acetone-H ₂ 0) . The eluate was lyophil- ized to obtain 80mg of the title compound.

IRfKBrJc ^"1 : 1800-1720

NMR(D ₂ 0) £ : 1.27 (6H, t, J=7.4Hz) , 2 , 30-2.80 (3H, s)

Example 6

Preparation of (IR, 5S, 6S) -2-[ (2S, 4S) -2-{ (carbamoyl-1- (hydro- xy) ethylcarbamoyl) methylmercaptomethyl}pyrrolidin-4-yl] thio- 6- [ (R) -l-hydroxyethyl ] -l-methyl-l-carbapen-2-em-3-carboxylic acid

(1) The reaction was carried out in the same manner a that of Example 1-(1) , starting from 580mg of 4-nitrobenzyl 2-diphenoxyphosphoryloxy-6-[ (R) carbapen-2-em-3-carboxy-late and 310mg of (2S,4S) -[1-(p nitrobenzyloxycarbonyl)-2-{ (carbamoyl-1-(hydroxy)ethylcarba moyl)methylmercaptomethyl}-4-mercapto]pyrrolidine to obtai 620mg of 4-nitrobenzyl (IR,5S,6S)-2-[ (2S,4S)-l-(p-nitrobenz yloxycarbonyl)-2-{ (carbamoyl-1-(hydroxy)ethylcarbamoyl)meth ylmercaptomethyl}pyrrolidin-4-yl]thio-6-[ (R)-1-hydroxyethy- l]-l-methyl-l-carbapen-2-em-3-carboxylate.

IRfNeatJcm ^"1 : 1765, 1725-1700, 1605 NMR(CDC1 ₃ ) $ : 1.1-1.7 (6H,m) , 5.0-5.6(4H,m) , 7.4-7.8(4H,m) , 8.21(4H,d,J=8.5Hz)

(2) The compound obtained in the above (1) was hydrog¬ enated in the same manner as that of Example l-(2) to obtai 170mg of the title compound.

Melting Point 176-177.5°C (dec.) IR(KBr)cm ^_1 1765, 1705-1675 NMR(D ₂ 0) S 1.21(3H,d,J=9Hz) , 1.27(3H,d,J=6Hz) ,

1.42-2.03 (2H,m) , 2.53-4.36(10H,m)

Example 7

Preparation of (IR,5S,6S)-2-[ (2S,4S)-2-{ (hydroxy-1-(hydroxy- methyl)ethylcarbamoyl)methylmercaptomethyl}pyrrolidin-4-yl] thio-6-[ (R)-l-hydroxyethyl]-l-methyl-l-carbapen-2-em-3-car- boxylic acid

(1) The reaction was carried out in the same manner as that of Example l-(l), starting from 720mg of 4-nitrobenzyl- 2-diphenoxyphosphoryloxy-6-[ (R)-l-hydroxyethyl]-1-methyl-l- carbapen-2-em-3-carboxylate and 310mg of (2S, S) -[1-(p-ni-

trobenzyloxycarbonyl) -2-{ (hydroxy-1- (hydroxymethyl) ethylcar¬ bamoyl) methy lmercaptomethyl }-4-mercapto] pyrrolidine to ob¬ tain 840mg of 4-nitrobenzyl (IR, 5S, 6S) -2-[ (2S,4S) -l-(p-ni- trobenzyloxycarbonyl) -2-{ (hydroxy-1- (hydroxymethyl) ethylcar- bamoyl) methylmercaptomethyl}pyrrolidin-4-yl] thio-6-[ (R) -1- hydroxyethyl]-l-methyl-l-carbapen-2-em-3 -carboxylate.

IR(Nujol)cm ^_1 : 1760, 1725-1710, 1705, 1605 NMR(CDC1 ₃ ) $ : 1.26(3H,d,J=9Hz) , 1.36 (3H,d, J=6Hz) , 5.15-5:45(6H,m) , 7.40-7.75 (6H,m) ,

8.25(6H,d,J=8Hz)

(2) The compound obtained in the above (1) was hydrog¬ enated in the same manner as that of Example l-(2) to obtain 225mg of the title compound.

Melting Point : 168-171°C (dec.)

IR(Nujol)cm ^-1 : 1750, 1725-1700, 1580

NMR(D ₂ 0) : 1.22 (3H,d,J=7Hz) , 1.28(3H,d,J=6Hz) , 1.6-1.9(2H,m) , 2.49-2.90(2H,m)

Example 8

Preparation of (IR, 5S, 6S) -2-[ (2S, 4S) -2-{ (carbamoyl-1- (hydro¬ xymethyl) ethylcarbamoyl) ethy lmercaptomethyl }pyrrolidin-4- yl]-thio-6-[ (R) -l-hydroxyethyl] -l-methyl-l-carbapen-2-em-3- carboxylic acid

(1) The reaction was carried out in the same manner as that of Example 1-(1), starting from 760mg of 4-nitrobenzyl- 2-diphenoxyphosphoryloxy-6-[ (R)-l-hydroxyethyl]-1-methy1-1- carbapen-2-em-3-carboxylate and 350mg of (2S,4S)-[1-(p-ni- trobenzyloxycarbonyl)-2-{ (carbamoy1-1-(hydroxymethyl)ethy1- carbamoyl}-4-mercapto]pyrrolidine to 840mg of 4-nitrobenzyl (IR,5S,6S) -2-[ (2S,4S)-l-(p-nitrobenzyloxycarbonyl)-2-{ (carb¬ amoy1-1-(hydroxymethyl)ethylcarbamoyl)ethylmercaptomethyl}-

pyrrolidin-4-yl] thio-6-[ (R) -l-hydroxyethyl] -1-methyl-l-car- bapen-2-em-3-carboxylate.

IR(Nujol)cm ^"1 : 1765, 1725, 1710, 1690 NMR(CDC1 ₃ ) £ : 1.23 (3H,d, J=9Hz) , 1.33 (3H, d, J=8Hz) ,

4.42-4.56(2H,m) , 4.82-5.5 (10H,m) , 7.4-7.70(4H,m) , 8.25 (6H, d, J=8Hz)

(2) The compound obtained in the above (1) was hydrog- enated in the same manner as that of Example l-(2) to obtain 250mg of the title compound.

Melting Point : 163-167°C (dec.) IR(KBr)cm ^-1 : 1750, 1725-1710 NMR(D ₂ 0) S ^: l-22(3H,d,J=7Hz) , 1.28 (3H,d,J=6Hz) ,

1.45-2.00(4H,m) , 2.46-2.95 (3H,m) ,

3.01-3.13(3H,m)

Example 9

Preparation of (IR, 5S, 6S) -2-[ (2S, 4S) -2-{ (2-cyano-l- (carbam¬ oylmethyl) ethylcarbamoyl) ethylmercaptomethyl}pyrrolidin-4- yl] thio- 6- [ (R) -l-hydroxyethyl ]-l-methyl- l-carbapen-2-em-3- carboxylic acid

(1) The reaction was carried out in the manner as that of Example 1-(1), starting from 820mg of 4-nitrobenzyl-2- diphenoxyphosphoryloxy-6-[ (R) -l-hydroxyethyl] -1-methyl-l-ca- rbapen-2-em-3-carboxylate and 380mg of (2S, 4S) -[l-(p-nitro- benzyloxycarbonyl) -2-{ (cyano-1- (carbamoylmethyl) ethylcarbam¬ oyl) ethy lmercaptomethyl}-4-mercapto]pyrrolidine to obtain 920mg of 4-nitrobenzyl ( IR, 5S , 6S) -2- [ (2S , 4S) -1- (p-nitroben- zyloxycarbonyl) -2{ (cyano-l-( carbamoylmethyl) ethylcarbamoyl) - ethylmercaptomethyl}pyrrolidin-4-yl]thio-6-[ (R) -l-hydroxyet¬ hyl] -l-methyl-l-carbapen-2-em-3-carboxylate .

IR(Nujol)cm ^_1 : 1765, 1715-1705, 1665 NMR(CDC1 ₃ ) S : 1.18 (3H,d,J=9Hz) , 1.26 (3H,d,J=9Hz) ,

5.15-5.47(6H,m) , 7.40-7.75 (6H,m) , 8.25(6H,d,J=8Hz)

(2) The compound obtained in the above (1) was hydrog¬ enated in the same manner as that of Example l-(2) to obtain 270mg of the title compound.

Melting Point : 163-164°C (dec.)

IR(Nujol)cm ^_1 : 1750, 1725-1705, 1665 NMR(CDC1 ₃ ) £ : 1.22 (3H,d,J=7Hz) , 1.28 (3H,d,J=6Hz) ,

1.60-1.90(2H,m) , 2.50-3.10(4H,m)

Example 10

Preparation of (IR, 5S, 6S) -2-[ (2S,4S) -2-{ (carbamoylethyl)mer- captomethyl}pyrrolidin-4-yl]thio-6-[ (R) -l-hydroxyethyl]-1- methyl-l-carbapen-2-em-3-carboxylic acid

(1) The reaction was carried out in the same manner as that of Example 1-(1) , starting from 520mg of 4-nitrobenzyl- 2-diphenoxyphosphoryloxy-6-[ (R) -l-hydroxyethyl]-1-methyl-l- carbapen-2-em-3-carboxylate and 270mg of (2S,4S) -[l-(p-nit- robenzyloxycarbonyl) -2{ (carbamoylethyl)mercaptomethyl}-4-me- rcapto]pyrrolidine to obtain 480mg of 4-nitrobenzyl (1R,5S, 6S) -2-[ (2S, 4S) -1-(p-nitrobenzyloxycarbonyl) -2-{ (carbamoylet- hyl)mercaptomethyl}pyrrolidin-4-yl]thio-6-[ (R) -l-hydroxyeth¬ yl]-l-methyl-l-carbapen-2-em-3-carboxylate.

IR NujolJcm ^"1 : 1775-1760, 1690-1660

NMR(CDC1 ₃ ) S : 1.26 (3H,d,J=7Hz) , 1.33 (3H,d,J=7Hz) , 1.70-2.15(2H,m) , 2.30-2.80 (2H,m) ,

4.50-4.83 (4H,m) , 7.40-7.65 (4H,m) ,

8.25(6H,d,J=8Hz)

(2) The compound obtained in the above (1) was hydrog¬ enated in the same manner as that of Example l-(2) to obtain 120mg of the title compound.

Melting Point : 162-163°C (dec.)

IR(KBr)cm ^_1 : 1765, 1715-1690, 1580 NMR(D ₂ 0) $ : 1.27 (3H,d,J=8Hz) , 1.33 (3H,d,J=8Hz) ,

2.17-2.50(4H,m) , 2.65-2.95 (2H,m)

Example 11

Preparation of (IR, 5S, 6S) -2-[ (2S, 4S) -2-{ (hydroxyethyl)mer- captomethyl}pyrrolidin-4-yl]thio-6-[ (R) -l-hydroxyethyl]-1- methyl-l-carbapen-?-em-3-carboxylic acid

(1) The reaction was carried out in the same manner as that of Example 1-(1) , starting from 480mg of 4-nitrobenzyl- 2-diphenoxyphosphoryloxy-6-[ (R) -l-hydroxyethyl]-1-methyl-l- carbapen-2-em-3-carboxylate and 210mg of (2S, 4S) -[1- (p- nitrobenzyloxycarbonyl) -2-{ (hydroxyethyl) -mercaptomethy}-4- mercapto]pyrrolidine to obtain 325mg of 4-nitrobenzyl(IR, 5S, 6S) -2-[ (2S, 4S) -1-(p-nitrobenzyloxycarbonyl) -2-{ (hydroxy- ethyl) mercaptomethyl}pyrrolidin-4-yl]thio-6-[ (R) -l-hydrox¬ yethyl]-l-methyl-l-carbapen-2-em-3-carboxylate.

IR(Neat)cm ^_1 : 3400, 1765-1725, 1610-1590 NMR(CDC1 ₃ ) : 1.20-1.80(6H,m) , 5.1-5.6(4H,m) , 7.3-7.70(4H,m) , 8.20 (4H,d,J=8Hz)

(2) The compound obtained in the above (1) was hydrog¬ enated in the same manner as that of Example l-(2) to obtain 80mg of the title compound.

Melting Point : 172-174°C (dec.)

IR(KBr)cm ^_:L : 1760-1730, 1595-1775

NMR(D ₂ 0) S : 1.21 (3H,d,J=8Hz) , 1.28 (3H,d,J=7Hz) ,

1.51-2.l(lH,m) , 2.99-2.9(lH,m)

Example 12

Preparation of (IR,5S,6S)-2-[ (2S,4S)-2-{ (4-(2-hydroxyethyl) piperazinyl)carbonylethylmercaptomethyl}pyrrolidin-4-yl] thio-6-[ (R) -l-hydroxyethyl]-l-methyl-l-carbapen-2-em-3- carboxylic acid

(1) The reaction was carried out in the same manner as that of Example 1-(1), starting from 720mg of 4-nitrobenzyl- 2-diphenoxyphosphoryloxy-6-{ (R)-l-hydroxyethyl]-1-methyl-l- carbapen-2-em-3-carboxylate and 250mg of (2S,4S)-[1-(p-ni- trobenzyloxycarbonyl)-2-{ (4-(2-hydroxyethyl)piperazinyl)car- bonyethylmercaptomethy}-4-mercapto]pyr o1idine to obtain 870 mg of 4-nitrobenzyl(IR,5S, 6S) -2-[ (2S,4S) -1-(p-nitrobenzy- loxycarbonyl) -2-{ (4-2-hydroxyethyl)piperazinyl)carbonylethy- lmercaptomethyl}pyrrolidin-4-yl]thio-6-[ (R)-l-hydroxyethyl]- l-methyl-l-carbapen-2-em-3-carboxylate.

IR(Neat)cm ^'1 : 3400, 1760-1750, 1690, 1525 NMR(CDC1 ₃ ) S - 1.26(3H,d,J=6Hz) , 1.33 (3H,d,J=6Hz) , 1.80-2.18(4H,m) , 2.27-2.38(4H,m) ,

5.10-5.63(4H,m) , 7.48(2H,d,J=8Hz) , 7.62(2H,d,J=8Hz) , 8.16-8.27(4H,d,J=8Hz)

(2) The compound obtained in the above (1) was hydrog- enated in the same manner as that of Example l-(2) to obtain

210mg of the title compound.

Melting Point : 168-170°C (dec.) IR(KBr)cm ^-1 : 3400, 1760-1735, 1600-1580 NMR(D ₂ 0) £ : 1.23 (3H,d,J=8Hz) , 1.28(3H,d,J=8Hz) ,

1.45-1.85(2H,m) , 2.30-2.66(3H,m) _; 2.88-3.0(3H,m)

Example 13

Preparation of (1R,5S,6S)-2-[ (2S,4S)-2-{ (4 (2-hydroxyet_ιyl) piperazinylcarbonylmethylcarbamoyl)ethylmercaptomethyl}pyrr- olidin-4-yl]thio-6-[ (R)-l-hydroxyethyl]-1-methyl-l-carbapen- 2-em-3-carboxylic acid

(1) The reaction was carried out in the same manner as that of Example 1-(1), starting from 680mg of 4-nitrobenzyl- 2-diphenoxyphosphoryloxy-6-[ (R)-l-hydroxyethyl]-1-methyl-l- carbapen-2-em-3-carboxylate and 290mg of (2S,4S)-[1-(p-ni- trobenzyloxycarbonyl)-2-{ ( (4-(2-hydroxyethyl)piperazinylcar- bonyImethylcarbamoyl)ethylmercaptomethy}-4-mercapto]-pyrro¬ lidine to obtain 580mg of 4-nitrobenzyl(IR,5S,6S)-2-[ (2S, 4S)-l-(p-nitrobenzyloxycarbonyl)-2-{ ( (4-(2-hydroxyethyl)pip¬ erazinylcarbonylmethylcarbamoyl)ethylmercaptomethyl}pyrroli- din-4-yl]thio-6-[ (R)-l-hydroxyethyl]-l-methyl-l-carbapen-2- em-3-carboxylate.

IR(Neat)cm ^_1 : 3400, 1770, 1705, 1650

NMR(CDC1 ₃ ) g : 1.25(3H,d,J=7.5Hz) , 1.32(3H,d,J=6Hz) ,

2.21-2.38(2H,m) , 3.10-4.83 (4H,m) , 4.85(2H,br. s) , 5.24(2H,s),

7.52(2H,d,J=8Hz) , 7.65(2H,d,J=8Hz) ,

8.25(4H,d,J=8Hz)

(2) The compound obtained in the above (1) was hydrog- enated in the same manner as that of Example l-(2) to obtain 90mg of the title compound.

Melting Point : 180-182°C (dec.) IR(KBr)cm ^_:L : 3400, 1755, 1680 NMR(D ₂ 0) 8 1•22 (3H,d,J=8Hz) , 1.30(3H,d,J=8Hz) ,

1.57-2.35(6H,m) , 3.01-3.55(2H,m)

Example 14

Preparation of (IR,5S,6S)-2-[ (2S,4S)-2-{ (aminocarbonyloxyme- thylcarbamoyl)ethylmercaptomethyl}pyrrolidin-4-yl]thio-6- [ (R)-l-hydroxyethyl]-l-methyl-l-carbapen-2-em-3-carboxylic acid

(1) The reaction was carried out in the same manner as that of Example 1-(1), starting from 420mg of 4-nitrobenzyl- 2-diphenoxyphosphoryloxy-6-[ (R)-l-hydroxyethyl]-1-methyl-l- carbapen-2-em-3-carboxylate and 280mg of (2S,4S) -[1-(p- nitrobenzyloxycarbonyl)-2-{ (aminocarbonyloxymethylcarbamoy- 1) -ethylmercaptomethy}-4-mercapto]pyrrolidine to obtain 380mg of 4-nitrobenzyl(IR,5S,6S)-2-[ (2S,4S)-1-(p-nitrobenzy- loxycarbonyl)-2-{ (aminocarbonyloxymethylcarbamoyl)ethylmer¬ captomethyl}-pyrr ^' olidin-4-yl]thio-6-[ (R)-l-hydroxyethyl]-1- methyl-l-carbapen-2-em-3-carboxylate.

IR(Neat)cm ^-1 : 3400, 1765, 1725-1715, 1610-1600, 1512,

1360 NMR(CDC1 ₃ ) δ : 1.1-1.6(6H,m) , 4.25-5.8(6H,m) ,

7.4-8.2(4H,m) , 8.15(4H,d,J=8.5Hz)

(2) The compound obtained in the above (1) was hydrog¬ enated in the same manner as that of Example l-(2) to obtain llOmg of the title compound.

Melting Point : > 170°C (dec.)

IR(KBr)cm ^_1 : 1750, 1700-1690, 1600-1580 NMR(D ₂ 0) £ : 1.23 (3H,d,J=7.0Hz) , 1.29(3H,d,J=6Hz) ,

1.6-2.1(2H,m) , 2.5-3.0(2H,m)

Example 15

Preparation of (IR,5S,6S)-2-[ (2S,4S)-2-{2-(ureidomethylcar-

bamoyl) ethylmercaptomethyl}pyrrolidin-4-yl]thio-6-[ (R) -1- hydroxyethyl]-l-methyl-l-carbapen-2-em-3-carboxylic acid

(l) The reaction was carried out in the same manner as that of Example 1-(1), starting from 580mg of 4-nitrobenzyl- 2-diphenoxyphosphoryloxy-6-[ (R) -l-hydroxyethyl]-1-methyl-l- carbapen-2-em-3-carboxylate and 275mg of (2S, 4S) -[1- (p- nitrobenzyloxycarbonyl) -2-{2-(ureidomethylcarbamoyl) ethylm- ercaptomethy}-4-mercapto]pyrrolidine to obtain 425mg of 4- nitrobenzyl (lR,5S,6S)-2-[(2S,4S) -l-(p-nitrobenzyloxycarbo- nyl) -2-{2-(ureidomethylcarbamoyl) ethylmercaptomethyl}pyrrol- idin-4-yl]thio-6-[ (R) -l-hydroxyethyl]-l-methyl-l-carbapen-2- em-3-carboxylate.

IR(Neat)cm ^_1 : 3400, 1775, 1710-1690, 1610, 1525-1510,

1350 NMR(CDC1 ₃ ) £ : 1.25-1.35 (6H,m) , 3.05-4.25 (10H,m) ,

4.80(2H,br. s) , 5.20(2H,s), 5.40(2H,dd,J=14Hz) , 7.56 (2H,d,J=8Hz) ,

7.68(2H,d,J=8Hz) , 8.26(4H,d,J=8Hz)

(2) The compound obtained in the above (1) was hydrog¬ enated in the same manner as that of Example l-(2) to obtain 90mg of the title compound.

Melting Point : > 175°C (dec.)

IR(KBr)cm ^"1 : 1760, 1710, 1650, 1580

NMR(D ₂ 0) : 1.21 (3H,d,J=9Hz) , 1.28 (3H,d,J=6Hz) , 1.42-2.10(4H,m) , 2.53-4.36 (14H,m)

Example 16

Preparation of (IR, 5S, 6S) -2-[ (2S, 4S) -2-{2- (aminomethylcar¬ bamoyl) ethylmercaptomethyl}pyrrolidin-4-yl]thio-6-[ (R) -1- hydroxyethyl]-l-methyl-l-carbapen-2-em-3-carboxylic acid

(1) The reaction was carried out in the same manner as that of Example 1-(1) , starting from 620mg of 4-nitrobenzyl- 2-diphenoxyphosphoryloxy-6-[ (R) -l-hydroxyethyl]-l-methyl-l- carbapen-2-em-3-carboxylate and 380mg of (2S, 4S) -[1-(p-ni- trobenzyloxycarbonyl) -2-{2-( (4-nitrobenzyloxycarbonyl)methy- lcarbamoyl) ethylmercaptomethyl}-4-mercapto]pyrrolidine to obtain 420mg of 4-nitrobenzyl (IR, 5S, 6S) -2-[ (2S, 4S)'-1- (p- nitrobenzyloxycarbony1) -2-{2-( (4-nitrobenzyloxycarbonyl)me- thylcarbamoyl) ethylmercaptomethyl}pyrrolidin-4-yl]thio-6- [ (R) -l-hydroxyethyl]-l-methyl-l-carbapen-2-em-3-carboxylate.

IR(Nujol)cm ^_1 : 1765-1700, 1710-1690, 1660-1650, 1530-1510

(2) The compound obtained in the above (1) was hydrog¬ enated in the same manner as that of Example l-(2) to obtain 190mg of the title compound.

Melting Point : > 180°C (dec.) IR(Nujol)cm ^_1 : 1760-1750, 1590-1580, 1350

NMR(D ₂ 0) g : 1.21 (3H,d,J=7Hz) , 1.30 (3H,d,J=7Hz) ,

1.40-2.1(4H,m) , 2.55-3.08(4H,m) , 3.12-4.35(9H,m)

Example 17

Preparation of (IR, 5S, 6S) -2-[ (2S, 4S) -2-{2-(methoxymethylcar¬ bamoyl) ethylmercaptomethyl}pyrrolidin-4-yl]thio-6-[ (R) -1- hydroxyethyl]-l-methyl-l-carbapen-2-em-3-carboxylic acid

(1) The reaction was carried out in the same manner as that of Example 1-(1) , starting from 510mg of 4-nitrobenzyl- 2-diphenoxyphosphoryloxy-6-[ (R) -l-hydroxyethyl]-1-methyl-l- carbapen-2-em-3-carboxylate and 250mg of (2S , 4S) - [ 1- (p- nitrobenzyloxycarbony1) -2-{2-(methoxymethylcarbamoyl) ethylm- ercaptomethy}-4-mercapto]pyrrolidine to obtain 420mg of 4-

nitrobenzyl (1R,5S, 6S) -2-[ (2S,4S) -1-(p-nitrobenzyloxycarbon- yl) -2-{2-(methoxymethylcarbamoyl) ethylmercaptomethyl}pyrrol- idin-4-yl]thio-6-[ (R) -l-hydroxyethyl]-l-methyl-l-carbapen-2- em-3-carboxylate.

IR(Nujθl)cm~ ¹ : 1750, 1700, 1685, 1516

NMR(CDC1 ₃ ) g : 1.28 (3H,d,J=7Hz) , 1.37 (3H,d,J=7Hz) ,

1.65-2.10 (3H,m) , 2.35-2.85 (2H,m) , ^' 2.94(3H,S), 5.25(4H,S), 5.40-5.75(2H,m) , 7.56 (2H,d,J=9Hz) ,

7.66(2H,d,J=9Hz) , 8.26 (4H,d,J=9Hz)

(2) The compound obtained in the above (1) was hydrog¬ enated in the same manner as that of Example l-(2) to obtain llOmg of the title compound.

Melting Point : > 178°C (dec.) IR(KBr)cm ^_1 : 1760-1750, 1590-1580, 1350 NMR(D ₂ 0) £ : 1.22 (3H,d,J=7Hz) , 1.28 (3H,d,J=6Hz) , 1.45-2.10(4H,m) , 2.24-2.95 (2H,m) ,

3.13(3H,s)

The compounds of the following Examples 18 to 27 can be prepared in the same manner as that of Example 1 using the corresponding starting materials.

Example 18

(1) (IR, 5R, 6S)-2-[ (2S, 4S) -l-(p-nitrobenzyloxycarbony-

1) -2-{2- ( (4-cyanoethylpiperazinyl) carbony Imethy lcarbamoyl) ethylmercaptomethyl}pyrrolidin-4-yl] thio-6- [ (R) -l-hydrox¬ yethyl] -l-methyl-l-carbapen-2-em-3-carboxylic acid

IRfNeatJc ^"1 :' 3600, 1765, 1710-1685, 1610, 1520

NMR(CDC1 ₃ ) ^' : 1.30-1.38 (6H, dd, J=18Hz) , 2.25-2.83 (7H,m) ,

3.18-3.50(3H,m) , 5.20-5.52 (4H,m) , 7.56-7.69(4H,m) , 8.28 (4H, , J=9Hz)

(2) (lR,5S,6S)-2-[ (2S,4S)-2-{2-( (4-cyanoethylpiperazi- nyl) carbonyImethylcarbamoyl) ethylmercaptomethyl}pyrrolidin- 4-yl]thio-6-[ (R) -l-hydroxyethyl]-l-methyl-l-carbapen-2-em-3- carboxylic acid

: 298.2nm

: 1755, 1710, 1585

: 1.22(3H,d,J=6Hz) , 1.31(3H,d,J=6Hz) , 1.75-2.35(5H,m)

(1) (IR, 5S, 6S) -2-[ (2S,4S) -l-(p-nitrobenzyloxycarbon- yl) -2-{2-( (4-p-nitrobenzyloxycarbonylaminoethylpiperazinyl) carbonyImethylcarbamoyl) ethylmercaptomethyl}pyrrolidin-4- yl]-thio-6-[ (R) -l-hydroxyethyl]-1-methyl-l-carbapen-2-em-3- carboxylic acid

IR(Neat)cm~ ¹ : 1765-1750, 1710, 1660-1640, 1530-1510 NMR(CDC1 ₃ ) g : 1.30 (3H,d,J=7Hz) , 1.38 (3H,d,J=7Hz) ,

1.75-2.10(3H,m) , 2.80-3.90 (10H,m) , 3.90-4.40(4H,m) , 5.20-5.50 (6H,m) , 7.55(4H,d,J=8Hz) , 7.66-8.25 (4H,dd)

(2) (lR,5S,6S)-2-[ (2S,4S)-2-{2-((4-aminoethylpiperazi- nyl) carbonylmethylcarbamoyl) ethylmercaptomethyl}pyrrolidin- 4-yl]-thio-6-[ (R) -l-hydroxyethyl]-l-methyl-l-carbapen-2-em- 3-carboxylic acid

Uv : 297.8nm

IR(Nujol)cm~ ¹ : 1750, 1590-1580

NMR(D ₂ 0) : 1.22 (5H,d,J=7Hz) , 1.30 (3H,d,J=7Hz) , 1.45-1.95(3H,m) , 2.55-3.08 (4H,m) ,

3.12-4.35(10H,m)

Example 20

(1) (IR, 5S, 6S) -2-[ (2S,4S) -1- (p-nitrobenzyloxycarbon- yl) -2-{2- (4- (2-methoxyethyl) piperazinyl) carbony Imethylcar bamoylethylmercaptomethyl}pyrrolidin-4-yl]-thio-6-[ (R) -1- hydroxyethyl]-l-methyl-l-carbapen-2-em-3-carboxylic acid

IR(Neat)cm ^_1 : 1765-1755, 1700, 1685, 1516, 1392 NMR(CDC1 ₃ ) 8 : 1.27 (3H, d, J=7Hz) , 1.34 (3H, d, J=7Hz) ,

2.76(4H,m), 3.30(3H,S), 5.21(2H, ^* S), 7.50-7.69(4H,m) , 8.28 (4H, d, J=9Hz)

(2) (lR,5S,6S)-2-[ (2S,4S)-2-{2-(4-(2-methoxyethyl)pi- perazinyl) carbonyImethylcarbamoylethylmercaptomethyl}pyrro- lidin-4-yl]-thio-6-[ (R) -l-hydroxyethyl]-1-methyl-l-carbapen- 2-em-3-carboxylic acid

1750, 1650, 1585, 1380

NMR(D ₂ 0) £ ^: l-19(3H,d,J=7Hz) , 1.27 (3H,d,J=6Hz) ,

2.72(3H,s) , 2.79-3.14(4H,m) , 3.30(3H,s), 4.05-4.20(4H,m)

Example 21

(1) (lR,5S,6S)-2-[(2S,4S) -1- (p-nitrobenzyloxycarbon- y1) -2-{2-(4-(2-carbamoyloxyethy1)piperazinylcarbonyl)methy1- carbamoylethylmercaptomethyl}pyrrolidin-4-yl]-thio-6-[ (R) -1- hydroxyethy1] -l-methyl-l-carbapen-2-em-3-carboxylic acid

IR(Neat)cm ^-1 : 1785, 1740, 1705, 1525, 1348 NMR(CDC1 ₃ ) = 1.35 (3H,d,J=6Hz) , 1.48 (3H,d,J=7Hz) , 1.95-2.05(2H,m) , 2.65-3.40 (3H,m) ,

5.22(2H,S) , 7.5-7.8(4H,dd,J=19Hz) , 8.20(4H,d,J=8Hz)

(2) (lR,5S,6S)-2-[ (2S,4S)-2-{2-(4-(2-carbamoyloxyeth- yl) piperazinylcarbonyl)methylcarbamoylethy lmercaptomethyl} pyrrolidin-4-yl]-thio-6-[ (R) -l-hydroxyethyl] -1-methy 1-1- carbapen-2-em-3-carboxylic acid

725-1710, 1600-1580 NMR(D ₂ 0) 8 : 1.22(3H,d,J=7Hz) , 1.28 (3H,d,J=6Hz) ,

1.6-1.9(4H,m) , 2.5-2.9(lH,m)

Example 22

(1) (IR,5S,6S)-2-[ (2S,4S)-1-(p-nitrobenzyloxycarbon- yl)-2-{2-(4-(2-ureidoethyl)piperazinyl)carbonylmethylcar- bamoyl)ethylmercaptomethyl}pyrrolidin-4-yl]-thio-6-[ (R)-1- hydroxyethyl]-l-methyl-l-carbapen-2-em-3-carboxylic acid

IR NujolJcirT ¹ : 1770, 1700, 1610, 1525, 1350 NMR(CDC1 ₃ ) ^: 1•27(3H,d,J=7Hz) , 1.32 (3H,d,J=6Hz) ,

2.85-3.90(llH,m) , 4.80(2H,br. s) , 5.25(2H,S), 5.37(2H,dd,J=18Hz) , 7.56-7.68(4H,dd,J=18Hz) , 8.26(4H,d,J=8Hz)

(2) (lR,5S,6S)-2-[ (2S,4S)-2-{2-(4-(2-ureidoethyl)pip¬ erazinyl)carbonyImethylcarbamoyl)ethylmercaptomethyl}pyrrol- idin-4-yl]-thio-6-[ (R)-l-hydroxyethyl]-1-methy1-1-carbapen- 2-em-3-carboxylic acid

^W mlxl ^: 298.5nm

IR(Nujol)cm ^-1 : 1755, 1700, 1650, 1580 NMR(CDC1 ₃ ) 8 : 1.25(3H,d,J=7Hz) , 1.33(3H,d,J=6Hz) ,

1.42-2.03(2H,m) , 2.53-4.50(45H,m)

Example 23

(1) (1R,5S, 6S)-2-[ (2S,4S)-l-(p-nitrobenzyloxycarbon-

yl) -2-{ (2-methoxymethyloxymethylcarbamoylethyl)mercaptometh- y 1 }pyrrolidin-4-yl ] -thio-6- [ (R) -l-hydroxyethyl ] -1-methy 1-1- carbapen-2-em-3-carboxylic acid

IRfNeatJcm ^"1 : 1765-1750, 1700, 1680

NMR(CDC1 ₃ ) 8 : 1.27(3H,d,J=7Hz) , 1.34 (3H, d, J=7Hz) ,

2.25-2.60(3H,m) , 3.35(3H,s), 5.21(2H,S), 7.55-7.70(4H,m) , 8.26 (4H,d, J=9Hz)

(2) (lR,5S,6S)-2-[ (2S,4S)-2-{ (2-methoxymethyloxymeth- ylcarbamoylethyl)mercaptomethyl}pyrrolidin-4-yl] -thio-6- [ (R) -l-hydroxyethyl] -l-methyl-l-carbapen-2-em-3-carboxylic acid

U max ^: 298.7nm

IR(Nujol)cm ^-1 : 1750-1745, 1700, 1655, 1580, 1350 NMR(D ₂ 0) ^: l-20(3H,d,J=7Hz) , 1.26(3H,d,J=7Hz) ,

2.73(3H,s), 2.77-3.15(4H,m) , 3.33(3H,s),

4.10-4.15(3H,m)

Example 24

(1) (IR, 5S, 6S) -2-[ (2S,4S) -1- (p-nitrobenzyloxycarbon- yl) -2-{ (2-methylcarbamoyloxymethylcarbamoylethyl)mercapto- methyl}pyrrolidin-4-yl]-thio-6-[ (R) -l-hydroxyethyl ] -1-meth- yl-l-carbapen-2-em-3 -carboxylic acid

IR(Neat)cm ^_1 : 3500-3400, 1765, 1720-1700, 1605 NMR(CDC1 ₃ ) 8 ^: 1- 1-1 • 8 (6H,m) , 5.10-5.75 (4H,m) ,

7.4-7.8 (4H,m) , 8.21(4H,d,J=8.5Hz)

(2) (lR,5S,6S)-2-[(2S,4S)-2-{(2-methylcarbamoyloxyme- thylcarbamoylethyl)mercaptomethyl}pyrrolidin-4-yl]-thio-6- [ (R) -l-hydroxyethyl ] -l-methyl-l-carbapen-2-em-3-carboxylic acid

UV^ ma2°x _: 297.8nm

63

IR(Nujol)cm -1 1750, J725-1705, 1580 NMR(D ₂ 0) 8 1.25(3H,d,J=7Hz) , 1.28(3H,d,J=8Hz) , 1.6-1.9(2H,m) , 2.4-2.9(2H,m)

Example 25

(1) (IR, 5S, 6S) -2-[ (2S, 4S) -1-(p-nitrobenzyloxycarbon- yl)-2-{ (2-ureidoethylcarbamoylethyl)mercaptomethyl}pyrroli- din-4-yl]-thio-6-[ (R) -l-hydroxyethyl]-l-methyl-l-carbapen-2- em-3-carboxylic acid

IR(NeatJem ^"1 : 1770-1760, 1710-1705, 1610-1605, 1525 NMR(CDC1 ₃ ) g : 1.27(3H,d,J=7Hz) , 1.33 (3H,d,J=7Hz) . 3.10-3.95(13H,m) , 4.85(2H,br. s) ,

5.24(2H,S), 5.50(2H,d,J=7Hz) , 7.56-7.68(4H,dd,J=18Hz) , 8.26(4H,d,J=8Hz)

(2) (IR, 5S, 6S) -2-[ (2S, 4S) -2-{ (2-ureidoethylcarbamoy- lethyl) mercaptomethy 1 }pyrrolidin-4-yl ] -thio-6- [ (R)-l- hydroxyethyl] -1-methy l-l-carbapen-2-em-3-carboxylic acid

IR(Nujol)cm ^"1 : 1755, 1650, 1580 NMR(D ₂ 0) g : 1.20 (3H,d, J=9Hz) , 1.25 (3H,d, J=6Hz) ,

1.45-2.10(4H _; m) , 2.53-4.35(14H,m)

Example 26

(1) (IR, 5S, 6S) -2-[-(2S, 4S) -1- (p-nitrobenzyloxycarbon- yl) -2-{ (2-(p-nitrobenzyloxycarbonyl)aminoethylcarbamoyleth- yl) _nercapto_nethyi}pyrrolidin-4-yl] -thio-6- [ (R) -l-hydrox¬ yethyl ]-l-methyl-l-carbapen-2-em-3 -carboxylic acid

IRΪNeatJcm ^"1 : 1765-1750, 1710, 1665-1650, 1510 NMR(CDC1 ₃ ) g : 1.24 (3H, d, J=7Hz) , 1.36 (3H,d, J=7Hz) ,

2.35-2.50(2H,m) , 3.15-3.46 (3H,m) ,

3.56-4.40(12H,m) , 5.12-5.50 (6H,m) , 7.36-7.80(6H,m) , 8.24 (6H, d, J=8Hz)

(2) (lR,5S,6S)-2-[(2S,4S)-2-{ (2-aminoethylcarbamoy- lethyl )mercaptomethyl}pyrrolidin-4-yl]-thio-6-[ (R) -l-hydrox¬ yethyl ] -l-_rιethyl-l-carbapen-2-em-3-carboxylic aciD

UVÏ€a : 298.5nm

IR(Nujol)cm ^_1 : 1770, 1705, 1610, 1525, 1350 NMR(D ₂ 0) ÏŠ 1.25(3H,d,J=7Hz) , 1.33 (3H,d,J=6Hz) ,

1.43-2.15(4H,m) , 2.55-4.25 (14H,m)

Example 27

(1) (1R,5S,6S)-2-[ (2S,4S)-l-(p-nitrobenzyloxycarbon- yl)-2-{ (2-methoxymethyloxyethylcarbamoylethyl)mercapto eth- yl}pyrrolidin-4-yl]-thio-6-[ (R)-l-hydroxyethyl]-1-methyl-l- carbapen-2-em-3-carboxylic acid

IR(Neat)cm ^-1 : 1770-1750, 1710-1700, 1690, 1605, 1520 NMR(CDC1 ₃ ) $ : 1.28(3H,d,J=7Hz) , 1.37(3H,d,J=7Hz) ,

1.65-2.10(5H,m) , 2.35-2.85(4H,m) , 2.94-3.10(4H,m) , 3.25(3H,s), 5.40-5.74(2H,m) , 7.56(2H,d,J=9Hz) ,

7.66(2H,d,J=9Hz) , 8.26(4H,d,J=9Hz)

(2) (lR,5S,6S)-2-[(2S,4S)-2-{(2-methoxymethyloxyethyl- carbamoylethyl)mercaptomethyl}pyrrolidin-4-yl]-thio-6-[ (R)- l-hydroxyethyl]-l-methyl-l-carbapen-2-em-3-carboxylic acid

50, 1700, 1590-1580, 1380 NMR(D ₂ 0) 8 : 1.22(3H,d,J=7Hz) , 1.28 (3H,d, J=7Hz) , 1.48-2.00(4H,m) , 2.46-2.95 (2H,m) ,

3.15(3H,S)

The specific examples of the compound of formula(I)

which can be prepared in the same manner as that of Example 1 are listed in the following Table.

66

70

CO NCH, o 2 r—

CON

' ^CH ₃ 0 2

CON N(CH2) ₂ OH o 2

CON » _* NCH - ^~ CN 2

CH ₂ CON^O o

NH, r ^~ \

CH ₂ CON NH >__ o r ^~ \

CH ₂ CON NCH ₃ o

CH ₂ C0N NCH,OH O

71

149 Me H H CH CH ₂ C0NH ₂ O 2

150 Me H H CH CH ₂ C0NHCH ₃ O 2

151 Me H H CH CH ₂ CON(CH ₃ ) ₂ O 2

152 Me H H CH CH ₂ OCONH ₂ O 2

153 Me H H CH CH ₂ NHCONH ₂ O 2

155 Me H H CH CH ₂ CON^ O 2

162 Me anion H CH CHjCON ⁽ . ^ ^{3 •} CH ₃ O 2

163 Me H H CH CH ₂ CON _^ NCH ₂ OH O 2

164 Me H H CH CH ₂ CON _t NCH ₂ NH ₂ O 2

165 Me H H CH CHoCON NCH ₂ CN O ₂

166 Me H H CH CH ₂ CON Vj(CH ₂ ) ₂ OH O 2

167 Me H H CH CHjCON N(CH ₂ ) ₂ NH ₂ O 2

187 Me H ^â–  C=NH CH CH ₂ CONH ₂ 0 2 CH ₃

188 Me H -C=NH CH CH ₂ OCONH ₂ 0 2 CH ₃

1 89 Me H -C=NH CH CH ₂ NHCONH ₂ 0 2 CH ₃

190 Me H H CN CH ₂ OH 0

1 91 Me H H CN CH ₂ CN 0

1 92 Me H H CN CH ₂ NH ₂ O

1 93 Me H H CN CH ₂ CONH ₂ 0

1 94 Me H H CN CH ₂ CONHCH ₃ O

195 Me H H CN CH ₂ CON(CH ₃ ) ₂ 0

1 96 Me H H CN CH ₂ OCONH ₂ O

1 97 Me H H CN CH ₂ NHCONH ₂ O

1 98 Me H H CN CH ₂ OH O 2

1 99 Me H H CN CH ₂ CN O 2 H CN CH ₂ NH ₂ O 2 H CN CH ₂ CONH ₂ O 2 H CN CH ₂ CON(CH ₃ ) ₂ O 2 H CN CH ₂ CON(CH ₃ ) ₂ O 2 H CM CH ₂ NHCONH ₂ O 2

H CN CH ₂ OCONH ₂ O 2

263 Me H H CH ₂ OH

264 Me H H CH ₂ OH

266 Me H H CH ₂ OH

272 Me anion H CH ₂ OH

273 Me H H CHoOH

274 Me H H CH ₂ OH

275 Me H H CH ₂ OH

276 Me H H CH ₂ OH 2

H H N NCH ₂ CH ₂ OH

H H N NCH ₂ CH ₂ CN

H H N NCH ₂ CH ₂ OCNH ₂ Q

H H N NCH ₂ CH ₂ OH o 2

H H N NCH ₂ CH ₂ CN o 2 O H H N _{% ^} NCH^^OCNHg o 2

H H N NCHoCH ₂ NH ₂ O 2 O H H N NCH ₂ CH ₂ CNH ₂ O 2

O H H N NCH ₂ CH ₂ CN(CH ₃ ) ₂ O 2

_^

H H N _< NCH ₂ CH ₂ <_.N(CH _a ) ₂ O 2

H

H

H

H

H

H

H

H

H

H

H H NH ₂ NH

H (CH3) ₂ N-J

H H MeNFt MH H H N(Et)a N-,

H NH,

H NHCH ₃

WCKate

t. H v,' ^; Et

H

H H

H H

H H

H ;-!

'H H

H H

H H

H H N NCH ₂ CH ₂ CN N-l 2

2

3

4

5

1

2

3

4

1

2

3

4

5

6

1

2

3

4

1

2

540 Me H ^â–  C=NH OCONH2 CH ₃ 541 Me H C=N-I OCONH ₂ CH,

98

Reference Example 1

44.6g of 2-amino-2-methyl-l-propanol, 65g of diethyl carbonate and 7g of potassium carbonate were mixed an stirred while heating at 120°C to 140°C, and then methano produced during the reaction was removed with a water trapping apparatus. The reaction solution was concentrate and then allowed to stand in a refrigerator to obtain white precipitate. The resulting product was dissolved i ethyl ether and filtered to remove the insoluble materials. The filtrate was concentrated and then allowed to stan under ice-cooling to obtain a white solid which is the filtered and dried to obtain 25.7g of the desired produc 4,4-dimethyl-oxazolidin-2-one.

Melting Point : 52-53°C

¹ H NMR(DCC1 ₃ ) δ : 1.35(s,6H,2CH ₃ ) , 4.00(s,2H,CH ₂ ) ,

Reference Example 2

11.5g of the compound prepared in Reference Example l was dissolved in 200ml of anhydrous tetrahydrofuran and the resulting solution was cooled to -50°C to -60°C and 70ml of n-butyl lithium was slowly added thereto under nitrogen atmosphere. At the same temperature 22g of 2-bromopropionyl bromide was added dropwise thereto under stirring. After stirring -for one hour at. the same temperature, the reaction mixture was slowly warmed to 0°C and then stirred for on hour. The reaction mixture was diluted with water and the extracted with ethyl ether. The organic layer was combined, dried and concentrated under reduced pressure to obtain 18 of the desired product 3-(2-bromopropionyl) -4,4-dimethyl- oxazolidin-2-one.

^ ^â–  H NMR(DCC1 ₃ ) & : 1.60(s,6H,2CH ₃ ) , 1.82(d, 3H,CH ₃ ) ,

4.05(s,2H,CH ₂ ) , 5.80(q,lH,CH,CH ₃ )

Reference Example 3

0.59g of azetinone and 0.5g of activated zinc were added to 40ml of anhydrous tetrahydrofuran. 1.06g of the compound prepared in Reference Example 2 was added dropwise thereto while heating under reflux and then the reaction mixture was heated for further 30 minutes under reflux. Saturated ammonium chloride solution was added to the mix¬ ture to complete the reaction. Ethyl acetate was added to the mixture to extract the reaction product and the extract¬ ed organic layers were combined, washed with distilled water, dried and concentrated under reduced pressure to obtain 470mg of the desired product (3S, 4S) -3- (lR-t-butyl- dimethylsilyloxyethyl)-4-[ (lR"-l-carboxyethyl) -2-(4"^"-di¬ methyl-2"-oxazolidinoyl) ]azetidin-2-one.

Melting Point : 171-172°C

¹ H NMR(DCC1 ₃ ) δ : 0.07(s, 3H) , 0.08(s,3H) , 0.87(s,9H) ,

1.19(d,3H,J=6Hz) , 1.21 (d, 3H,J=6Hz) , 1.58(Ξ,6H) , 3.01-3.04(s,lH) , 3.90-3.95 (m,lH) , 4.14-4.23(m,2H) , 6.00(s,lH,NH)

Reference Example 4

5.1g of the compound prepared in Reference Example 3 was dissolved in 200ml of tetrahydrofuran and then 3.5ml of 30% hydrogen peroxide waε added thereto. Then, lN-sodium hydroxide solution was slowly added dropwise to the reaction mixture with stirring at 5 to 10°C. The whole reaction mixture was stirred for 30 minutes at the same temperature, evaporated under reduced pressure to remove tetrahydrofuran solvent and extracted with ethyl acetate. The extract was adjusted to pH 3 to 4 by adding 10% hydrochloric acid εolu-

tion and evaporated under reduced pressure to remove th solvent to obtain 2.6g of the white desired product (3S,4S) 3-[ (lR-t-butyldimethylsilyloxyethyl) -4-(lR-1-carboxyethyl) ] azetidin-2-one.

Melting Point : 148.5-151.5°C

¹ H NMR(CDC1 ₃ ) $ : 0.07-0.08(s,6H) , 0.89(s,9H), 1.19

(d,3H,J=6Hz) , 1.27(d,3H,J=7Hz) , 2.78-2.74 (m,lH) , 3.05(d,1H,J=4.4Hz,2H) , 3.98(d,lH,J=4.82Hz) , 4.25-4.17(m,2H) ,

6.50(s,lH,NH)

Reference Example 5

0.6g of the compound prepared in Reference Example was dissolved in 5ml of anhydrous acetonitrile and th resulting solution was stirred for 30 minutes. Then 2.Og o magnesium p-nitrobenzylmalonate was added thereto and th mixture was stirred for 18 hours at 65°C. The reactio mixture was evaporated to remove the solvent and the residu was suspended in 50ml of ethyl acetate and then washe successively with lN-hydrochloric acid solution, 10% potas sium carbonate and saline. After removing the solvent, th residue was purified with column chromatography to obtai 70mg of the desired product (3S,4S) -3-[ (R) -1-(t-butyldimeth ylsilyloxy)ethyl]-4-[ (R) -1-(p-nitrobenzylacetoxy)carboxyet- hyl]azetidin-2-one.

^λ H NMR(CDC1 ₃ ) : 0.07 (s,6H,CH3X2) , 0.9(s,9H), 1.10(d,3H,

CH ₃ ), 1.23(d,3H,CHX3) , 2.29(dd,lH), 2.9 (m,lH) , 3.66(s,2H), 3.96(dd,lH), 4.20 (m,lH), 5.39(s,2H) , 7.56 and 8.25(d, 2H, 0 ) .

Reference Example 6

2.87g of the compound prepared in Reference Example 5 was dissolved in 30ml of methanol and 3ml of 6N-hydrochloric acid solution was added thereto. The reaction mixture was stirred for 2 hours at room temperature and then adjusted to pH 7 to 8 with 0. IN-phosphate buffer and 10% potassium carbonate. After removing the solvent from the reaction mixture, the residue was extracted with ethyl acetate. The extract was washed with water and concentrated under reduced pressure. The residue was purified with column chromatogra- phy to obtain 1.7g of the desired product (3S, 4S) -3-[ (R) -1- (t-butyldimethylsilyloxy) ethyl]-4-[ (R) -1-(p-nitrobenzylace- toxy)carboxyethyl]azetidin-2-one.

¹ H NMR(CDC1 ₃ ) S : 1.30(d,6H), 2.90(dd, 1H,CH) , 2.94(q,lH, CH) , 3.65-3.70(ABq,2H,CH ₂ ) , 3.84(dd,lH,

CH) , 4.15(m,lH,CH) , 7.50 and 8.25(d,

Reference Example 7

0.4g of the compound prepared in Reference Example 6 was dissolved in 5ml of anhydrous acetonitrile and to the resulting solution were added 7.4g of tosyl azide and 0.2g of triethylamine. The reaction solution was stirred for 2 hours at room temperature and then concentrated under re¬ duced pressure. The residue was subjected to column chroma¬ tography to obtain 400mg of the desired product (3S,4S)-3- [ (R) -l-hydroxyethyl]-4-[ (R) -l-methyl-3-diazo-3- (p-nitroben- zyloxycarbonyl) -2-oxopropyl]azetidin-2-one.

^ ^• H NMR(CDC1 ₃ ) ξ : 1.25(d,6H), 2.95(dd,lH), 3.77(q,lH,

CH) , 3.86(dd,lH,CH) , 4.15 (m, 1H,CH) , 5.38 (s,2H,CH ₂ ) , 7.55 and 8.30(d,2H, 0)

Reference Example 8

0.4g of the compound prepared in Reference Example was added to 20ml of the mixed solvent of anhydrous ethy acetate and anhydrous hexane and the resulting mixture wa heated for one hour under reflux. A catalytic amount o rhodium acetate was added thereto under warming. The reac tion mixture was stirred under reflux for one hour and the filtered with diatomaceous earth to remove the solvent t obtain 380mg of the desired product p-nitrobenzy1- (1R, 5S, 6S) -2-keto-6-[ (IR) -l-hydroxyethyl]-1-methyl-l-carba- pen-2-em-3-carboxylate.

^ ^• H NMR(CDC1 ₃ ) ^' : 1.23 (d, 3H, S-methyl) , 1.40(d,3H,

CH ₃ CHOH) , 2.86(q,lH,CH) , 3.30(dd, 1H,CH) , 4.30(dd,lH,CH) , 4.36(q,lH,CH) , 4.78(s, 1H,CH), 5.28 and 5.42 (ABq,2H,CH ₂ ) ,

7.58 and 8.30(d,2H, 0 )

Reference Example 9

13.2g of trans-4-hydroxy-L-proline was dissolved in 111ml of 2N-sodium hydroxide solution, and 23.76g of p- nitrobenzyloxycarbonylchloroformate dissolved in 20ml of methylene chloride was added thereto while stirring under ice-cooling and the mixture was stirred for 2 hours at the same temperature. Then 50ml of 2N-sodium hydroxide was added to the mixture to separate the layers. The organic layer was removed, and 18.5g of concentrated sulfuric acid was added to the aqueous layer to precipitate the product. The resulting product was filtered, washed with distilled water and dried to obtain the desired product trans-l-(p- nitrobenzyloxycarbonyl) -4-hydroxy-L-proline in the yield of 83%.

Melting Point : 132-135°C

IR(Nujol)cm ^"1 : 3300, 1738, 1660, 1665, 1520

Reference Example 10

120g of the compound prepared in Reference Example was dissolved in 50ml of methanol and the resulting solutio was added dropwise to 50ml of tetrahydrofuran. To thi mixture was added dropwise 60ml of ethyl ether solution o diazo ethane under nitrogen atmosphere while stirring wit ice-cooling and the whole mixture was stirred for 20 hours. The reaction solution was concentrated under reduced pres sure to obtain the desired product trans-1-(p-nitrobenzy loxycarbonyl)-4-hydroxy-L-proline methyl ester in the yiel of 70%.

IR(Neat)cm ^"1 : 1748, 1695, 1518, 1438, 1360, 1250, 1175

Reference Example 11

5.18g of the compound prepared in Reference Example 10, 3.79g of trimethylamine and 3.77g of t-butyldimethylsily chloride were dissolved in 50ml of anhydrous di ethylforma mide and the resulting solution was stirred for 3 hours a room temperature, diluted with distilled water and the extracted with ethyl acetate. The organic layer was sepa¬ rated and washed successively with distilled water, 1.0N- hydrochloric acid solution and saline, dried and concentrat¬ ed under reduced pressure to obtain the desired produc trans-l-[ (p-nitrobenzyloxycarbonyl)-4-(4-(t-butyldi- ethyl silyloxy) ]-L-proline methyl ester in the yield of 68%.

[ σ. ] _p ⁹ = -36.2° (C=1.00 CHCl ₃ )

IR(Neat)cm ^_1 : 1750, 1710, 1517, 1415, 1355, 1250, 1115 NMR(CDC1 ₃ ) ξ : 0.08(9H,s), 1.8-2.4 (2H,m) , 3.3-3.8 (2H,m) , 3.63(3H,S), 3.72(3H,ε), 5.20(1H,J=14Hz) ,

5.23(lH,s), 7.42(2H,d,J=9Hz) , 8.15(2H,d,J=9Hz)

Reference Example 12

5.64g of the compound prepared in Reference Example 11 was dissolved in 60ml of anhydrous tetrahydrofuran and l.Olg of sodium borohydride and 3.52g of calcium chloride were added to the resulting solution. The reaction solution was refluxed under heating for one hour and diluted with dis¬ tilled water and then extracted with ethyl acetate. The organic layer was separated, washed with distilled water and saline, dried and concentrated under reduced pressure to obtain the desired product (2S,4S)-[1-(p-nitrobenzyloxycar- bonyl)-2-(hydroxymethyl)-4-(t-butyldimethylsilyloxy) ]pyrro- lidine in the yield of 75%.

[ o. ] 'D ⁹ = -40.1° (C=1.00 CHC1 ₃ ) IR(Neat)cm ^_1 : 1670, 1504, 1420, 1405, 1240, 1100

NMR(CDC1 ₃ ) £ : 0.07(6H,s), 0.87(9H,s), 1.4-2.1(2H,m) ,

3.38-3.84(4H,m) , 3.9-4.5(2H,m) , 5.22(2H,s), 7.47(2H,d,J=9Hz) ,

8.17(2H,d,J=9HZ)

Reference Example 13

32. 6g of the compound prepared in Reference Example 12 was dissolved in 64ml of anhydrous pyridine and 28g of p- toluenesulfonyl chloride was added to the resulting solution and then the mixture was stirred for 12 hours at room tem- perature . The reaction solution was diluted with distilled water and then extracted with ethyl acetate . The organic layer was separated , washed with saline , 1 . ON hydrochloric acid solution and distilled water , dried and concentrated under reduced pressure to obta in the des ired product ( 2S , 4S) - [ l- (p-nitrobenzy loxycarbony 1) -2- (p-toluenesulf ony lo- xy ethyl ) -4 - ( t-butyldimethylsi lyloxy) ] pyrrolidine in the yield of 75% .

106

IR(Neat)cm -1 1700, 1518, 1342, 1265, 1172, 1090

Reference Example 14

35.8g of the compound prepared in Reference Example 13 and 19g of sodium iodide were added to 90ml of ethyle- thylketone and the mixture waε refluxed under heating. The reaction εolution was cooled to room temperature and then filtered. The filtrate was concentrated under reduced pressure and the residue was dissolved in ethyl acetate, washed with each of distilled water and saline, dried and concentrated under reduced pressure to obtain the desired product (2S,4S)-[1-(p-nitrobenzyloxycarbonyl) -2-(iodometh- yl)-4-(t-butyldimethylsilyloxy) ]-pyrrolidine in the yield of 70%.

Melting Point : 88-92°C

IR(Neat)cm ^_1 : 1700, 1512, 1405, 1353, 1248 NMR(CDC1 ₃ ) £ : 0.07(6H,ε), 0.87(9H,s), 1.4-2.5(2H,m) , 3.1-

3.8(4H,m), 3.95-4.38(2H,m) , 5.22(2H,s), 7.5-8.3(4H,d,J=9Hz)

Reference Example 15-A

5g of. the compound prepared in Reference Example 14 was dissolved in 50ml of dimethyIformamide, and then 2g of 3- mercaptopropionate ethyl ester, 1.7g of calcium iodide and 1.7g of triethylamine were added to the resulting solution while stirring. The reaction mexture was warmed to 60°C to 70°C and allowed to react for 4 hours. The reaction mixture waε cooled to room temperature, diluted with ethyl acetate, washed several times with distilled water and then washed with IN hydrochloric acid solution and saline, dried and concentrated under reduced pressure to obtain the desired product (2S,4S) -[1-(p-nitrobenzyloxycarbony1)-2-(ethyloxy- carbonylethylthiomethyl)-4-(t-butyldimethylsilyloxy) ]pyrrol-

idine in the yield of 60%.

IR(Neat)cm ^~1 : 1745, 1705, 1695, 1510, 1405, 1342 NMR(CDC1 ₃ ) $ : 0.07(6H,ε), 0..9(9H,ε), 1.55-2.5(4H,m) , 3.0(3H,ε), 3.15-3.8(4H,m) , 3.95-4.25(4H, m) , 5.22(2H,s), 7.25-8.35(9H,d,J=9Hz)

Reference Example 15-B

llg of the compound prepared in Reference Example 15-A was dissolved in 20ml of methanol, and 30ml of 2N-sodium hydroxide solution was added to the resulting solution while stirring under ice-cooling. The reaction mixture was stirred for 30 minutes, adjusted to neutral pH value by adding lN-acetic acid and then evaporated under reduced preεεure to remove the solvent. The residue was extracted with ethyl acetate. The extract was dried, concentrated under reduced pressure and then subjected to column chroma- tography to obtain the desired product (2S, 4S) - [ 1- (p- nitrobenzyloxycarbonyl)-2-(hydroxycarbonylethylthiomethyl)- 4-(t-butyldimethylsilyloxy) ]pyrrolidine in the yield of 85%.

IR(Neat)cm ^_1 : 3500, 1725, 1690, 1425, 1350 NMR(CDC1 ₃ ) $ : 0.08(6H,s), 0.87(9H,s), 1.45-2.20(4H,m) ,

3.10-3.50(4H,m) , 5.22(2H,S), 7.25-8.5(4H, d,J=8HZ)-

Reference Example 15-C

4g of the compound prepared in Reference Example 15-B was dissolved in 40ml of anhydrous acetonitrile and 1.56g of carbonyldii idazole was added to the resulting solution under ice-cooling. The mixture was stirred for 30 minutes and 2.7g of triethylamine and 1.06g of glycinamide hydro¬ chloride were added thereto. The whole mixture was stirred for 2 hours under ice-cooling and then for one hour at room

temperature. The reaction solution was diluted with ethy acetate, washed succesεively with distilled water, 1N hydrochloric acid solution, 5% sodium bicarbonate solutio and saline, dried and then concentrated under reduced preε sure. The residue was subjected to column chromatograph eluting with ethyl acetate:n-hexane (1:1) to obtain th desired product (2S,4S) -[1-(p-nitrobenzyloxy.carbonyl)-2- (carbamoylmethylcarbamoylethylthiomethyl)-4-(t-butyldimeth¬ ylsilyloxy) ]-pyrrolidine in the yield of 65%.

IR(Neat)cm ^-1 : 1725, 1710, 1695, 1515, 1420, 1325 NMR(CDC1 ₃ ) 8 : 0.07(6H,s), 0.8(9H,s), 1.25-1.45(2H,m) ,

2.25-2.50(4H,m) , 3.95-4.25(2H,m) , 5.25 (2H,ε), 7.5-8.5(4H,d,J=8Hz)

Reference Example 15-D

2.93g of the compound prepared in Reference Example 15- C was dissolved in 15ml of methanol and 2ml of 6N- hydrochloric acid εolution waε added to the resulting εolu- tion while εtirring under ice-cooling. The mixture was stirred for 2 hourε at the εame temperature. The reaction solution was concentrated under reduced presεure and the reεidue waε extracted with ethyl acetate. The extract waε dried and then concentrated under reduced pressure to remove the solvent. The residue was subjected to column chromatog¬ raphy eluting with ethyl acetate:n-hexane (3:1) to obtain the desired product (2S,4S)-[l-(p-nitrobenzyloxycarbonyl)-2- (carbamoylmethylcarbamoylethylthiomethyl)-4-(hydroxy) ]pyrro- lidine in the yield of 93.3%.

IR(Neat)cm ^-1 : 3600, 1720, 1700, 1690, 1510, 1420 NMR(CDC1 ₃ ) S : 1.80-2.18(2H,m) , 2.65-3.05(2H,m) , 3.09(2H, s) , 3.30-3.35(2H,m) , 3.85-4.50(2H,m) , 4.96-

5.24(2H,d,J=8Hz) , 7.66-8,26(4H,d,J=8Hz)

Reference Example 15-E

2.17g of the compound prepared in Reference Example 15- D was dissolved in 30ml of anhydrous dichloromethane and 0.59 of methanesulfonyl chloride and 0.71g of triethylamine were added to the resulting solution while εtirring under ice-cooling. Then, the mixture waε εtirred for 2 hourε at the same temperature. The reaction solution was then washed with distilled water, IN hydrochloric acid solution and saline, dried and concentrated under reduced pressure to obtain the desired product (2S,4S)-[l-(p-nitrobenzyloxycar- bonyl)-2-(carbamoylmethylcarbamoylethylthiomethyl)-4-(mesy- loxy) ]pyrrolidine in the yield of 66.1%.

IR(Neat)cm~ ¹ : 1710, 1700, 1690, 1510, 1435, 1350, 1050 NMR(CDC1 ₃ ) £ : 2.05-2.60(3H,m) , 3.03(3H,s), 5.25(2H,s),

7.53(2H, ^" d,J=8Hz) , 8.25(2H,d,J=8Hz)

Reference Example 15-F

To 20ml of di ethylformamide were added 2g of the compound prepared in Reference Example 15-E and 0.57g of potaεsium thioacetate and then the reaction mixture was stirred for 4 hourε at 70 to 80°C. The reaction mixture was cooled to room temperature, diluted with ethyl acetate and then waεhed εeveral times with diεtilled water, dried and concentrated under reduced pressure. The residue was εub- jected to column chromatography eluting with ethyl acetate to obtain the deεired product (2S,4S)-[l-(p-nitrobenzyloxy- carbonyl) -2- (carbamoylmethylcarbamoylethylthiomethyl) -4- (acetylthio) ]pyrrolidine in the yield of 65.2%.

IR(Neat)cm ^_1 : 1725, 1710, 1690, 1510, 1420, 1350

NMR(CDC1 ₃ ) 8 : 2.40-3.15(4H,m) , 3.25(3H,ε), 3.75-4.50

(4H,m) , 5.23(2H,ε), 7.25-8.23 (4H,d,J=8Hz)

Reference Example 15-G

1.5g of the compound prepared in Reference Example 15-F waε dissolved in 15ml of methanol and 0.2g of sodium ethox- ide was added to the resulting εolution while stirring under ice-cooling. The mixture was stirred for 15 minutes at the same temperature. The reaction mixture was added to acetic acid to neutralize and then concentrated under reduced presεure. The residue was extracted with ethyl acetate. The extract waε dried and then concentrated under reduced preεsure to obtain the desired product (2S, 4S) -1- (p- nitrobenzyloxycarbonyl)-2-(carbamoylmethylcarbamoylethylthi- omethyl)-4-mercaptopyrrolidine in the yield of 85.2%.

IR(Neat)cm ^_1 : 1710, 1690, 1510, 1425

NMR(CDC1 ₃ ) δ 1.75-1.95(3H,m) , 2.45-2.85(2H,m) , 2.90-

3.15(2H,m), 3.21(2H,s), 3.25-4.30(4H, ) , 5.24(2H,ε) , 7.55-8.27(4H, ,J=8Hz)

Reference Example 16-A

The reaction was carried out in the same manner as that of Reference Example 15-A, using 7g of the compound prepared in Reference Example 14, 1.8ml of ethylthioglyconate, 2.7g of calcium iodide and 2.1ml of triethylamine to obtain the desired product (2S,4S)-[1-(p-nitrobenzyloxycarbonyl)-2-(et- hyloxycarbonylmethylthiomethyl)-4-(t-butyldimethylεilyloxy) ] -pyrrolidine in the yield of 75%.

IRfNeatJcm ^"1 : 1745, 1700, 1690, 1515, 1410 NMR(CDC1 ₃ ) £ : 0.07(6H,ε), 0.9(9H,s), 1.50-1.98 (2H,m) ,

2.02-2.25(2H,m) , 3.0(3H,s), 3.15-3.8(2H, m) , 3.95-4.25(4H,m) , 5.22(2H,s), 7.25-

8.35(4H,d,J=8Hz)

Reference Example 16-B

3g of the compound prepared in Reference Example 16- waε treated in the εame manner aε that of Reference Exampl 15-B to obtain the desired product (2S, 4S) -[1-(p-nitroben zyloxycarbonyl) -2-(hydroxycarbonyImethylthiomethyl) -4-(t-bu tyldimethylsilyloxy) ]pyrrolidine in the yield of 60%.

IRfNeatJcm ^"1 : 3600, 1720, 1695, 1520, 1420

NMR(CDC1 ₃ ) $ : 0.08(6H,m), 0.85(9H,s), 1.45-2.20(2H,m) ,

3.10-3.45(4H,m) , 5.22(2H,s), 7.25-8.5 (4H,d,J=8Hz)

Reference Example 16-C

5g of the compound prepared in Reference Example 16-B was dissolved in 30ml of anhydrous acetonitrile solution an 2g of carbonyldiimidazole was added to the resulting solu tion while stirring under ice-cooling. After stirring th mixture, 3. Og of triethylamine and 1.5g of glycinamid hydrochloride were succeεsively added thereto and the reac tion mixture was then treated in the same manner aε that o Reference Example 15-C to obtain the deεired product (2S, 4S) -[1-(p-nitrobenzyloxycarbony1) -2-(carbamoylmethylcarbamo ylmethylthio ethyl) -4-(t-butyldimethylεilyloxy) ]pyrrolidine in the yield of 80%.

IR(Neat)cm ^_1 : 1720, 1705, 1695, 1570

NMR(CDC1 ₃ ) J ^' : 0.06(6H,s), 0.86(9H,ε), 1.88-2.22 (2H,m) ,

3.22(2H,S) , 5.25(2H,S), 7.53 (2H,d,J=8Hz) ,

8.25(2H,d,J=8Hz)

Reference Example 16-D

3.2g of the compound prepared in Reference Example 16- waε treated in the same manner aε that of Reference Exampl 15-C to obtain the deεired product (2S,4S)-[1-(p-nitrobenzy loxycarbonylj-2-(carbamoylmethylcarbamoylmethylthiomethyl)- 4-(hydroxy) ]pyrrolidine in the yield of 85.2%.

IR(Neat)cm ^_1 : 3600, 1715, 1700, 1510, 1400 NMR(CDC1 ₃ ) : 1.80-2.10(2H,m) , 2.65-3.05(2H,m) , 3.09(2H,

S) , 3.30-3.55(2H,m) , 3.85-4.50(2H,m) , 4.95-5.24 ^' (2H,ε) , 7.66-8.26(4H,d,J=8Hz)

Reference Example 16-E

2g of the compound prepared in Reference Example 16- waε dissolved in 30ml of anhydrous dichloromethane, an 0.75g of triethylamine and 0.6g of methanesulfonyl chloride were added to the reεulting εolution. Then, the reaction mixture waε treated in the εame manner as that of Reference Example 15-E to obtain the deεired product (2S,4S) -[1-(p- nitrobenzyloxycarbony1)-2-(carbamoyImethylcarbamoyImethylth- iomethyl)-4-(mesyloxy) ]pyrrolidine in the yield of 72%.

IR(Neat)cm ^"1 : 1715, 1700, 1695, 1500, 1410, 1355 NMR(CDC1 ₃ ) g : 2.05-2.60(3H,m) , 3.03(3H,s) , 5.25(2H,s),

7.53-8.25(4H,d,J=8Hz)

Reference Example 16-F

1.9g of the compound prepared in Reference Example 16-E waε added to 15ml of dimethyIformamide and 0.5g of potaεεium thioacetate and the resulting mixture was treated in the same manner as that of Reference Example 15-E to obtain the desired product (2S,4S)-[l-(p-nitrobenzyloxycarbonyl)-2-(ca¬ rbamoyImethylcarbamoylmethylthiomethyl)-4-(acetylthio) ]pyrr¬ olidine in the yield of 57%.

IR(Neat)cm ^-1 : 1725, 1750, 1695, 1500

NMR(CDC1 ₃ ) S = 2.40-3.15 (4H,m) , 3.21(3H,s) , 3.75-4.50 (5H, m) , 5.23(2H,ε), 7.59-8.23 (4H,d,J=8Hz)

Reference Example 16-G

2.Og of the compound prepared in Reference Example 16-F waε dissolved in 18ml of methanol and 0.25g of sodium me- thoxide was added to the resulting solution while stirring under ice-cooling. The reaction mixture was then treated in the same manner as that of Reference Example 15-G to obtain the desired product (2S,4S) -[1-(p-nitrobenzyloxycarbonyl) -2- (carbamoylmethylcarbamoylmethylthiomethyl) -4- (mercapto) ]py- rrolidine in the yield of 78%.

IR(Neat)cm ^_1 : 1710, 1695, 1520, 1425, 1350

NMR(CDCl ₃ ) 8 : 1-75-1.95(3H,m) , 2.45-2.85 (2H,m) , 2.90-3.15

(2H,m), 3.21(2H,s), 3.25-3.50 (2H,m) , 3.85- 4.30(2H,m), 5.24(2H,ε), 7.55-8.27 (4H,d,

J=8Hz)

Reference Example 17-A

4g of the compound prepared in Reference Example 14 was dissolved in 40ml of anhydrous acetonitrile and 1.56g of carbonyldiimidazole was added to the resulting εolution. After εtirring the mixture for 30 minuteε, 2.7g of triethy- lamine and 2.3g of aminoacetonitrile hydrochloride were added thereto and the whole mixture waε εtirred for 2 hours under ice-cooling and then one hour at room temperature. The reaction solution was diluted with ethyl acetate, washed sucpesεively with distilled water, lN-hydrochloric acid solution, 5% sodium bicarbonate solution and saline, dried and then concentrated under reduced pressure. The residue was subjected to column chromatography eluting with ethyl acetate: n-hexane (2:1) to obtain the desired product

(2S, 4S) -[ l-(p-nitrobenzyloxycarbonyl) -2- (cyanomethylcarbamo ylethylthiomethyl) -4- (t-butyldimethylsilyloxy) ]pyrrolidin in the yield of 65%.

IR(Neat)cm ^_1 : 2250, 1725-1710, 1690-1665 NMR(CDC1 ₃ ) g : 0.05(6H,s), 0.86(9H,s), 1.80-2.15 (4H,m) ,

5.25(2H,ε) , 7.25-8.21(4H,d,J=8Hz)

Reference Example 17-B

3.5g of the compound prepared in Reference Example 17- was dissolved in 15ml of methanol and 2ml of 6N-hydrochlori acid solution was added to the reεulting solution whil stirring under ice-cooling. The reaction solution wa stirred for 2 hours at the same temperature and concentrate under reduced presεure. The reεidue waε extracted wit ethyl acetate and the extract was dried and then concentrat ed under reduced preεsure to remove the solvent. The residu was subjected to column chromatography eluting with ethy acetate : n-hexane (3:1) to obtain the desired produc (2S, S) -[1-(p-nitrobenzyloxycarbonyl) -2-(cyanomethylcarbamo ylethylthio ethyl) -4-(hydroxy) ]pyrrolidine in the yield o 87.5%.

IR(Neat)cm ^_1 : 2250, 1710, 1695-1675, 1610, 1525 NMR(CDC1 ₃ ) : 1.52-1.95(2H,m) , 2.75-3.50(2H,m) , 4.05-

4.75(4H,m), 5.23(2H,S) , .7.53-8.22 (4H,d,

J=8HZ)

Reference Example 17-C

2.78g of the compound prepared in Reference Example 17 B was dissolved in 30ml of anhydrous dichloromethane an

0.62g of methanesulfonyl- chloride and 1.4g of triethylamin were added to the resulting solution while stirring unde ice-cooling. The mixture was stirred for 2 hours at th

same temperature. The reaction solution was waεhed with distilled water, lN-hydrochloric acid solution and saline, dried and then concentrated under reduced presεure to obtain the deεired product (2S,4S) -[1-(p-nitrobenzyloxycarbonyl) -2- (cyanomethylcarbamoylethylthiomethyl)-4-(meεyloxy) ]pyrroli- dine in the yield of 72.5%.

IR(Neat)cm ^_1 : 2250, 1710, 1700, 1690, 1510, 1435, 1350,

1050 NMR(CDC1 ₃ ) δ : 2.05-2.80(4H,m) , 3.03(3H,m), 7.53-8.25

(4H,d,J=8Hz)

Reference Example 17-D

2.8g of the compound prepared in Reference Example 17-C waε added to 20ml of dimethylformamide and then 0.65g of potaεεium thioacetate waε added thereto. The reaction mixture waε εtirred for 4 hourε at 70 to 80°C and then cooled to room temperature, diluted with ethyl acetate, waεhed εeveral times with diεtilled water, dried and concen¬ trated under reduced pressure. The residue was subjected to column chromatography eluting with ethyl acetate to obtain the desired product (2S,4S)-[1-(p-nitrobenzyloxycarbonyl)-2- (cyanomethylcarbamoylethylthiomethyl)-4-(acetylthio) ]pyrro lidine in the yield of 73.5%.

NMR(CDC1 ₃ ) 8 : 1.75-2.40(4H,m) , 3.25-4.55(6H,m) , 7.85-8.10

(4H,d,J=8Hz)

Reference Example 17-E

0.9g of the compound prepared in Reference Example 17-D was dissolved in 15ml of methanol, and O.lg of sodium me- thoxide waε added to the resulting solution while stirring under ice-cooling. The reaction solution was stirred for 15 minutes at the same temperature, neutralized with acetic

acid and then concentrated under reduced presεure. Th reεidue waε extracted with ethyl acetate and the extract wa dried and concentrated under reduced preεsure to obtain th desired product (2S,4S)-1- (p-nitrobenzyloxycarbonyl)-2-(cya nomethylcarbamoyiethylthiomethyl) -4-mercaptopyrrolidine i the yield of 63.8%.

IR(Neat)cm ^"1 : 1720-1690, 1605, 1530-1515

NMR(CDC1 ₃ ) f : 1.65-1.94 (3H,m) , 2.45-2.85 (2H,m) , 2.90-3.3 (4H,m), 3.25-3.50(2H,m) , 5.24(2H,s) , 7.55-

8.24(4H,d,J=8Hz)

Reference Example 18-A

6g of the compound prepared in Reference Example 14 wa dissolved in 40ml of anhydrous acetonitrile and 2.5g o carbonyldiimidazole waε added to the reεulting solutio while stirring under ice-cooling. After stirring the mix ture, 3.5g of triethylamine and 2. Ig of aminoacetonitril hydrochloride were succeεεively added thereto. Then, th subsequent reaction was carried out in the same manner a that of Reference Example 15-C to obtain the deεired produc (2S,4S) -[1-(p-nitrobenzyloxycarbonyl) -2-(cyanomethylcarbamo ylmethylthiomethyl)-4-(t-butyldimethylsilyloxy) ]pyrrolidine in the yield of 88.5%.

IR(Neat)cm ^_:L : 2250, 1735-1710, 1695-1665, 1610 NMR(CDC1 ₃ ) : 0.06(6H,s), 0.85(9H,s), 1.75-2.10 (4H,m) , 5.25(2H,s), 7.30-8.56 (4H,d,J=8Hz)

Reference Example 18-B

2.5g of the compound prepared in Reference Example 18- waε treated in the same manner as that of Reference Exampl 15-D to obtain the deεired product (2S, 4S) -[1-(p-nitrobenzy loxycarbonyl) -2-(cyanomethylcarbamoylmethylthiomethyl) -4-

(hydroxy) ]pyrrolidine in the yield of 90.8%.

IR(Neat)cm ^_1 : 2250, 1745-1710, 1690-1675 NMR(CDC1 ₃ ) : 1.75-2.25 (2H,m) , 2.55-3.10 (2H,m) , 3.90(2H,S), 3.30-3.45(3H,m) ,

4.95-5.24 (2H,s) , 7.66-8.25 (4H,d,J=8Hz)

Reference Example 18-C

2.5g of the compound prepared in Reference Example 18-B was dissolved in 30ml of anhydrous dichloromethane and 0.8g of methanesulfonyl chloride and l.Og of triethylamine were added to the resulting solution while εtirring under ice- cooling. The reεulting reaction mixture was treated in the same manner as that of Reference Example 15-E to obtain the desired product (2S, 4S) -[ 1- (p-nitrobenzyloxycarbonyl) -2- (cyanomethylcarbamoylmethylthiomethyl)-4-(mesyloxy) ]pyrroli- dine in the yield of 72%.

IR(Neat)cm ^_1 : 2250, 1725-1690, 1660, 1610 NMR(CDC1 ₃ ) £ : 2.10-2.45 (3H,m) , 3.03(3H,s) , 5.25(2H,s) ,

7.53-8.25(4H,d,J=8Hz)

Reference Example 18-D

2.5g of the compound prepared in Reference Example 18-C was added to 15ml of dimethylformamide and 0.8g of potaεεium thioacetate and then the reaction mixture was treated in the same manner as that of Reference Example 15-E to obtain the deεired product (2S, 4S) -[1- (p-nitrobenzyloxycarbonyl) -2- (cyanomethylcarbamoylmethylthiomethyl)-4-(acetylthio) ]pyrro- lidine in the yield of 63.5%.

IR(Neat)cm -1 2250, 1725, 1750, 1695, 1500 NMR(CDC1 ₃ ) 2.15-3.00(4H,m) , 3.2(3H,s) , 3.75-4.50 (5H,m) , 5.23(2H,ε) , 7.59-8.23 (4H,d,J=8Hz)

Reference Example 18-E

1.8g of the compound prepared in Reference Example 18-D was dissolved in 18ml of methanol, and 0.25g of sodium ethoxide was added to the resulting solution while εtirring under ice-cooling. The reaction mixture was treated in the same manner aε that of Reference Example 15-G to obtain the desired product (2S,4S).-[1-(p-nitrobenzyloxycarbonyl)-2- (cyanomethylcarbamoyImethylthiomethyl)-4-(mercapto) ]pyrroli- dine in the yield of 65.8%.

IR(Neat)cm ^-1 : 2250, 1735-1710, 1695-1675, 1610 NMR(CDC1 ₃ ) £ : 1.65-1.75(3H,m) , 2.45-2.85(2H,m) , 2.90-

3.30(5H,m), 3.50(2H,m), 5.24(2H,s), 7.55-8.27(4H,d,J=8Hz)

Reference Example 19-A

52g of the compound prepared in Reference Example 14 and 18.3g of potaεεium thioacetate were added to 50ml of dimethylformamide and the mixture waε warmed to 60 to 75°C and εtirred for 2 hourε.- The reaction εolution was cooled to room temperature, diluted with distilled water and ex¬ tracted with ethyl acetate. The organic layer waε waεhed with diεtilled water, εaline and 5% εodium bicarbonate εolution, dried and concentrated under reduced preεεure to obtain the desired product (2S,4S)-[1-(p-nitrobenzyloxycar¬ bonyl)-2-(acetylthiomethyl)-4-(t-butyldimethylsilyloxy) pyr¬ rolidine in the yield of 55%.

IR(Neat)cm~ ¹ : 1710-1700, 1610, 1530 NMR(CDC1 ₃ ) : 0.06(6H,s), 1.84(9H,S), 2.35(3H,s), 5.26

(2H,ε), 7.54-8.22 (4H,d,J=8Hz) ^'

Reference Example 19-B

5g of the compound prepared in Reference Example 19- was dissolved in 10ml of methanol and 2ml of 28% sodium methoxide waε added to the reεulting εolution while stirring under ice-cooling. The mixture was stirred for 15 minutes under nitrogen atmosphere and then lg of 2-iodoethanol dissolved in 10ml of methanol was added dropwiεe thereto at the same temperature. The reaction mixture was stirred for 4 hours at the same temperature and concentrated under reduced pressure to remove the solvent. The residue was dissolved in ethyl acetate, washed with distilled water, dried and then concentrated under reduced presεure to remove the solvent to obtain the desired product (2S,4S) -[1-(p- nitrobenzyloxycarbonyl)-2-(hydroxyethylthiomethyl)-4-(t-but ¬ yldimethylsilyloxy) ]pyrrolidine in the yield of 50%.

IR(NeatJem ^-1 : 1710, 1690-1675, 1610, 1525 NMR(CDC1 ₃ ) £ : 0.06(6H,s), 0.86(9H,ε), 1.88-2.22 (2H,m) ,

3.22(2H,S) , 5.25(2H,s), 7.53-8.29 (4H,d,J=8Hz)

Reference Example 19-C

850mg of the compound prepared in Reference Example 19- B and l l of 6N-hydrochloric acid εolution were added to 8ml of methanol and the mixture was stirred for one hour, dilut¬ ed with ethyl acetate, washed with distilled water, dried and then concentrated under reduced pressure. The residue was purified with column chromatography to obtain the de¬ sired product (2S,4S)-[1-(p-nitrobenzyloxycarbonyl)-2-(ace- toxyethylthiomethyl)-4-(hydroxy) ]pyrrolidine in the yield of 70%.

IR(CHCl ₃ )cm ^_1 : 1710, 1610, 1525

NMR(DMSO-dg) £ : 1.80-2.15 (2H, ) , 2.65-3.05 (2H, ) , 3.09

(2H,S) , 3.30-3.55(2H,m) , 5.24(2H,s) , 7.66-8.26(2H,d,J=8Hz)

Reference Example 19-D

1.5g of the compound prepared in Reference Example 19- C, 2.2g of triphenylphoεphine and 5g of diethylazodicarboxy- late were respectively added to 20ml of anhydrous tetrahy¬ drofuran solution, and the mixture was stirred for 2 hours under ice-cooling. 1.2g of thiolacetic acid was added to the reaction mixture at the same temperature and the mixture was stirred for 2 hours. The reaction solution was in- creased to room temperature and then εtirred for 20 hours, evaporated under reduced preεεure, diluted with ethyl ace¬ tate, waεhed with distilled water, dried and then concen¬ trated under reduced pressure. The residue was purified with column chromatography to obtain the desired product (2S,4S) -[1-(p-nitrobenzyloxycarbonyl) -2-(acetoxyethylthio- methyl)-4-(acetylthio) ]pyrrolidine in the yield of 70%.

IR(Neat)cm ^-1 : 1745, 1710, 1600, 1510, 1398, 1360, 1098 NMR(CDC1 ₃ ) S = 1•75-1.95(3H,m) , 2.45-2.85(lH,m) , 2.90- 3.15(2H,m), 3.21(2H,ε), 3.25-3.50(2H,m) ,

3.58-4.30(2H,m) , 5.24(2H,ε), 7.55-8.27 (4H,d,J=8Hz)

Reference Example 20-A

5g of the compound prepared in Reference Example 19-A waε diεεolved in 10ml of methanol, and 2ml of 28% sodium methoxide was added to the resulting solution under ice- cooling. The mixture was stirred for 15 minutes under nitrogen atmosphere. To this mixture waε added lg of 2- iodoethanol diεεolved in 10ml of methanol at the εa e tem¬ perature. The reaction mixture was εtirred for 4 hours at

the same temperature and concentrated under reduced pressure to remove the solvent. The residue waε diεεolved in ethyl acetate, waεhed with diεtilled water, dried and concentrated under reduced preεεure to remove the εolvent to obtain the desired product (2S, 4S) -[ 1- (p-nitrobenzyloxycarbonyl) -2- (hydroxyethylthiomethyl)-4-(t-butyldimethylsilyloxy) ]pyrro- lidine in the yield of 50%.

IR(Neat)cm ^_1 : 3600, 1740, 1690, 1523, 1400, 1345 NMR(CDC1 ₃ ) S : 0.06(6H,s) , 0.86(9H,ε), 1.51-2.1 (2H,m) ,

2.4-2.9(2H,m) , 3.22(2H,ε), 5.25(2H,s), 7.53-8.25(4H,d,J=8Hz)

Reference Example 20-B

1.5g of the compound prepared in Reference Example 20-A waε dissolved in 15ml of anhydrous pyridine and 0.8ml of acetic anhydride and 0.3g of dimethylaminopyridine were added to the reεulting εolution. The mixture was stirred for one hour at room temperature, diluted with ethyl ace¬ tate, washed with distilled water and then concentrated under reduced preεεure. The reεidue was purified with column chromatography to.obtain the desired product (2S,4S)- [1-(p-nitrobenzyloxycarbonyl) -2-(acetoxyethylthiomethyl) -4- (t-butyldimethylsilyloxy) ]pyrrolidine in the yield of 70%.

IR(Neat)cm ^-1 : 1740, 1725-1710, 1690, 1610 NMR(CDC1 ₃ ) S : 0.06(6H,s), 0.8(9H,s), 1.52-1.95(2H,m) , 2.4-2.65(2H,m) , 3.15-3.35 (3H,s) , 5.25

(2H,S), 7.53-8.25(4H,d,J=8Hz)

Reference Example 20-C

850mg of the compound prepared in Reference Example 20- B and 1ml of 6N-hydrochloric acid solution were -added to 8ml of methanol. The reaction mixture was stirred for one hour

at room temperature, diluted with 20ml of ethyl acetate waεhed with diεtilled water, dried and concentrated unde reduced preεεure. The reεidue waε purified with colum chromatography to obtain the deεired product (2S, 4S) -[1-( nitrobenzyloxycarbonyl) -2- (acetoxyethylthiomethyl) -4- (hy¬ droxy) ]pyrrolidine in the yield of 70%.

IR(Neat)cm ^_1 : 3600, 1720, 1680, 1510, 1410, 1342, 1225 NMR(CDC1 ₃ ) δ : 1.52-1.95 (2H,m) , 2.75-2.95 (2H,m)., 3.35 (3H,ε) , 4.05-4.75(2H,m) , 5.25(2H,s) , 7.53-

8.22(4H,d,J=8Hz)

Reference Example 20-D

1.5g of the compound prepared in Reference Example 20 C, 2.2g of triphenylphosphine and 5g of diethylazodicarboxy late were added to 20ml of anhydrous tetrahydrofuran solu tion. The mixture was stirred for 2 hours under ice cooling, and l.2g of thiolacetic acid was added thereto a the same temperature. The reaction mixture waε increased t room temperature and then εtirred for 20 hours, evaporate under reduced preεεure, diluted with ethyl acetate, waεhe with diεtilled water and then concentrated under reduce pressure. The residue was purified with column chromatogra phy to obtain the desired product (2S,4S) -[1- (p-nitrobenzy loxycarbonyl) -2- (acetoxyethylthiomethyl) -4- (acetylthio) ]- pyrrolidine in the yield of 70%.

IR(Neat)cm ^-1 : 1745, 1710, 1600, 1510, 1398, 1360, 1098

Reference Example 20-E

300mg of the compound prepared .in Reference Example 20 D was dissolved in 5ml of methanol and 2ml of IN sodiu hydroxide εolution waε added to the reεulting solution. Th mixture waε εtirred for 20 minuteε in nitrogen atmoεpher

under ice-cooling and then 2ml of IN hydrochloric aci solution was added thereto. The reaction solution wa concentrated under reduced pressure, and the residue wa dissolved in ethyl acetate, washed with diεtilled water an εaline, dried, and then concentrated under reduced preεεur to obtain an oily- residue. This oily reεidue waε purifie with column chromatography to obtain the deεired produc (2S, 4S) -[ 1- (p-nitrobenzyloxycarbonyl) -2- (hydroxyethylthio- methyl) -4-(mercapto) ]pyrrolidine in the yield of 46%.

IRfNeatJcm ^"1 : 3600, 1725, 1680, 1522, 1433, 1410, 1350 NMR(CDC1 ₃ ) £ : 1.85-2.22 (2H,m) , 2.65-3.05 (2H,m) ,

3.09(2H,m) , 3.30-3.55(2H,m) , 3.85-4.50 (2H,m) , 5.24(2H,s), 7.66-8.26 (4H,d,J=8Hz)

Reference Example 21-A

4g of the compound ^" prepared in Reference Example 15-B was disεolved in 50ml of anhydrous acetonitrile and 1.56g of carbonyldiimidazole was added to the resulting solution under ice-cooling. The reaction mixture was stirred for 30 minutes and 2.5g of triethylamine and 2g of 4- aminoacetyloxyethylpiperazine were added thereto. The whole mixture was stirred for 2 hours under ice-cooling and then for 4 hours at room temperature to complete the reaction. The reaction εolution waε diluted with ethyl acetate, waεhed with diεtilled water, lN-hydrochloric acid solution, 5% sodium bicarbonate solution and saline, dried and then con- centrated under reduced presεure. The residue was εubjected to column chromatography eluting with ethyl acetate:n-hexane (1:3) to obtain the deεired product (2S, 4S) -[1-(p-nitroben¬ zyloxycarbonyl) -2-{ (acetyloxyethylpiperazinylcarbamoyl) ethy- lmercaptomethyl}-4-(t-butyldimethylεilyloxy) Jpyrrolidine in the yield of 68%.

IR(Neat)cm ^-1 : 1740, 1710-1690, 1550

NMR(CDC1 ₃ ) § : 0.07(6H,s) , 0.8(9H,s), 1.25-1.55 (4H,m) ,

2.10-2.50(4H,m) , 5.25(2H,s) , 7.5-8.5 (4H,d,J=8Hz)

Reference Example 21-B

3.15g of the compound prepared in Reference Example 21 A waε diεεolved in 15ml of methanol, and 1.5ml of 6N-hydro chloric acid solution was added to the resulting εolutio while εtirring under ice-cooling. The reaction εolution wa stirred for 2 hours at the same temperature and then concen trated under reduced pressure. The residue was extracte with ethyl acetate, dried and then concentrated under re duced preεεure to remove the εolvent. The reεidue was subjected to column chromatography eluting with ethyl ace- tate:n-hexan (3:1) to obtain the desired product (2S,4S)-[1- (p-nitrobenzyloxycarbonyl) -2-{ (acetyloxyethylpiperazinyl) ca- rbamoylethylthiόmethyl}-4-(hydroxyJ ]pyrrolidine in the yield of 87.2%.

IR(Neat)cm~ ¹ : 3600, 1720, 1690, 1420

NMR(CDC1 ₃ ) S : 1.80-2.10 (2H,m) , 2.65-3.05 (4H,m) , 3.90

(2H,ε), 3.30-3.35 (2H,m) , 4.96-5.29 (2H,d,J=8Hz) , 7.66-8.26(4H,d,J=8Hz)

Reference Example 21-C

1.85g of the compound prepared in Reference Example 21- B waε diεεolved in 30ml of anhydrouε dichloromethane, and 0.45g of ethaneεulfonyl chloride and 0.62g of triethylamine were added to the resulting εolution. The reaction εolution waε εtirred for 2 hourε at the same temperature, waεhed with distilled water, lN-hydrochloric acid solution and saline, dried and then concentrated under reduced pressure to obtain the desired product (2S,4S) -[1- (p-nitrobenzyloxycarbonyl) -2- { (acetyloxypiperazinylcarbamoyl) ethylthiomethyl}-4-(mesy- loxy) ]pyrrolidine in the _, yield of 68.5%.

IR(Neat)cm ^-1 : 1745, 1710-1695, 1610, 1580

NMR(CDC1 ₃ ) = 2.10-2.55(4H,m) , 3.30-3.33 (3H,m) , 5.25

(2H,s), 7.58(2H,d,J=8Hz) , 8.25(2H,d,J=8Hz)

Reference Example 21-D

2g of the compound prepared in Reference Example 21-C was added to 20ml of dimethyIformamide, and 0.57g of potaε- sium thioacetate was added thereto. The reaction mixture

4was stirred for 4 hourε at 70 to 80°C and then cooled to room temperature. Then, the mixture was diluted with-ethyl acetate, washed several times with distilled water, dried and then concentrated under reduced pressure. The residue was subjected to column chromatography eluting with ethyl acetate to obtain the desired product (2S,4S)-[1-(p-nitro- benzyl- oxycarbόnyl)-2-{ (acetyloxyethylpiperazinyl)carbamoy- lethylthiomethyl}-4-(acetylthio) ]pyrrolidine in the yield of 65.2%.

IR(Neat)cm ^-1 : 1725, 1710, 1690, 1510, 1420, 1350 NMR(CDC1 ₃ ) g : 2.55-3.15(4H,m) , 3.25-3.35(3H,m) , 3.75-

4.50(4H,m), 5.23(2H,s), 7.25-8.55 (4H,d,J=8Hz)

Reference Example 21-E

2.Og of the compound prepared in Reference Example 21-D wad dissolved in 15ml of methanol, and 0.21g of sodium me- thoxide was added to the resulting solution while stirring under ice-cooling. The mixture was stirred for 15 minutes at the same temperature and then neutralized with acetic acid. The reaction solution was concentrated under reduced preεsure. The residue was extracted with ethyl acetate, dried and concentrated under reduced preεεure to obtain the deεired product (2S, 4S) -[ 1- (p-nitrobenzyloxycarbonyl) -2-

[ (hydroxyethylpiperazinylcarbamoyl) ethylthiomethyl]-4-merca- topyrrolidine in the yield of 70.1%.

IRfNeatJcm ^"1 : 1740, 1690, 1550 NMR(CDCl ₃ ) : 1.75-1.88 (3H,m) , 2.45-2.85(2H,m) , 2.90-

3.15(2H,m), 3.21(2H,S), 3.25-4.30(4H,m) , 5.24(2H,S) , 7.55-8.27(4H,d,J=8Hz)

Reference Example 22-A

3.5g of the compound prepared in Reference Example 15-B was disεolved in 50ml of anhydrouε acetonitrile, and 1.8g of carbonyldii idazole waε added to the resulting solution under ice-cooling. The mixture was stirred for 30 minutes and then 21g of triethylamine and 2.5g of glycina ide hydro¬ chloride were added thereto. The reaction mixture was stirred for 2 hourε under ice-cooling and then 11 hours at room temperature. The reaction solution was diluted with ethyl acetate, washed with distilled water, IN-hydrochloric acid solution, 5% sodium bicarbonate solution and saline, respectively, dried and then concentrated under reduced presεure. The reεidue was subjected to column chromatogra¬ phy eluting with ethyl acetate: n-hexane (10:1) to obtain the desired product (2S,4S) -[1-(p-nitrobenzyloxycarbonyl) -2- { (hydroxycarbonylmethylcarbamoyl)methylcarbamoylethylthiome- thyl}-4-(t-butyldimethylεilyloxy) ]pyrrolidine in the yield of 65%..

IR(Neat)cm ^_1 : 2250, 1725-1710, 1690-1665

NMR(CDC1 ₃ ) : 0.07(6H,s), 0.09(9H,s), 1.27-1.31(4H,m) ,

2.25-2.60(4H,m) , 3.95-4.45(2H,m) , 5.25 (2H,s) , ^" 7.5-8.25(4H,d,J=8Hz)

Reference Example 22-B

4.2g of the compound prepared in Reference Example 22-A

waε diεsolved in 50ml of anhydrous acetonitrile εolution, and 2.5g of carbonyldiimidazole waε added to the resulting solution under ice-cooling. The mixture was stirred for 30 minutes. To this mixture were added 21g of triethylamine and 2.06g of 4-aminohydroxyethylpiperazine, and the whole mixture was stirred for 2 hourε under ice-cooling and then 10 hours at room temperature. The reaction solution was diluted with ethyl acetate, washed with distilled water, IN hydrochloric acid solution, 5% sodium bicarbonate solution and saline, respectively, dried and then concentrated under reduced pressure. The residue was subjected to column chromatography eluting with ethyl acetate to obtain the desired product (2S, 4S) -[1-(p-nitrobenzyloxycarbonyl) -2- { (hydroxyethylpiperazinylcarbamoylmethylcarbamoyl)ethylthio- methyl}-4-(t-butyldimethylsilyloxy) ]pyrrolidine in the yield of 52%.

IR(Neat)cm ^_1 : 1730, 1710, 1680, 1510

NMR(CDC1 ₃ ) £ : 0.07(6H,s), 0.09(9H,s), 1.25-1.65(4H,m) , 2.25-2.45(4H,m) , 3.01-3.75(4H,m) , 4.15-

4.25(2H,m), 5.25(2H,S), 7.55-8.52 (4H,d, J=8Hz)

Reference Example 22-C

To 10ml of acetic anhydride were added 2.6g of the compound prepared in Reference Example 22-B and 2.5g of triethylamine and the mixture was stirred for 5 hours under reflux. The reaction solution was evaporated under reduced pressure. The reεidue waε diεεolved in ethyl acetate, waεhed εucceεsively with IN-hydrochloric acid εolution, 5% sodium bicarbonate solution and distilled water, dried and then distilled under reduced pressure. The residue was εubjected to column chromatography eluting with ethyl ace¬ tate to obtain the desired product ( 2S , 4S) - [ 1- (p- nitrobenzyloxycarbonyl)-2-{ (acetyloxyethylpiperazinylcarbam- oylmethylcarbamoyl)ethylthiomethyl}-4-(hydroxy) ]pyrrolidine

in the yield of 78.2%.

IRfNeatJcm ^"1 : 1725, 1710-1690, 1610, 1580 NMR(CDC1 ₃ ) ^' : 0.07(6H,s), 0.09(9H,S), 1.25-1.30 (2H,m) , 1.85-2.20(6H,m) , 4.15-4.50 (4H,m) , 3.77-

3.90(2H,m), 4.55-4.50(2H,m) , 5.24 (2H,d,J=6Hz) , 7.65-8.25(4H,d,J=8Hz)

Reference Example 22-D

2.55g of the compound prepared in Reference Example 22- C waε dissolved in 30ml of anhydrous dichloromethane, and 0.61g of ethanesulfonyl chloride and 0.71g of triethylamine were added to the reεulting εolution while εtirring under ice-cooling. The mixture waε then stirred for 2 hours at the same temperature. The reaction solution waε washed with distilled ^' water, IN hydrochloric acid solution and εaline, dried and concentrated under reduced preεεure to obtain the desired product (2S,4S)-[1-(p-nitrobenzyloxycar¬ bonyl)-2-{ (acetyloxypiperazinylcarbamoylmethylcarbamoyl)eth- ylthiomethyl}-4- (mesyloxy) ]pyrrolidine in the yield of 80.5%.

IR(Neat)cm ^_1 : 1710, 1690, 1510

NMR(CDC1 ₃ ) ^' : 2.10-2.45(6H,m) , 3.03(3H,m), 5.25(2H,s),

7.53-8.24(4H,d,J=8Hz)

Reference Example 22-E

• To 20ml of dimethylf ormamide was added 1.5g of the compound prepared in Reference Example 22-D and then 0.45g of potassium thioacetate was added thereto. The reaction mixture waε stirred for 4 hourε at 70 to 80°C, cooled to room temperature, diluted with ethyl acetate, washed several times with distilled water, dried and then concentrated under reduced pressure. The residue was subjected to column

chromatography eluting with ethyl acetate to obtain the desired product (2S, 4S) -[ 1- (p-nitrobenzyloxycarbonyl) -2- { (acetyloxyethylpiperazinylcarbamoylmethylcarbamoyl) ethylth- iomethyl}-4-(acetylthio) ]pyrrolidine in the yield of 65.9%.

IRfNeatJcm ^"1 : 1725, 1690, 1420, 1350

NMR(CDC1 ₃ ) : 2.15-3.00(6H,m) , 3.21(2H,m), 3.75-4.50

(4H,m), 5.23(2H,S), 7.59-8.23 (4H,d,J=8-Hz)

Reference Example 22-F

2.5g of the compound prepared in Reference Example 22-E waε dissolved in 18ml of methanol, and 0.31g of sodium methoxide was added to the resulting solution while stirring under ice-cooling. The reaction mixture was then treated in the same manner aε that .of Reference Example 15-G to obtain the deεired product (2S,4S) -[1-(p-nitrobenzyloxycarbonyl) -2- { (hydroxyethylpiperazinylcarbamoylmethylcarbamoyl) ethylthi- omethyl}-4-(mercapto) ]pyrrolidine in the yield of 60.1%.

IR(Neat)cm ^"1 : 1710, 1695, 1520, 1425, 1350 NMR(CDC1 ₃ ) g : 1.65-1.75 (3H,m) , 2.45-2.85 (4H,m) , 2.90-

3.30(5H,m) , 3.30(2H,ε) , 5.24(2H,ε), 7.55- 8.25(4H,d,J=8Hz)

Reference Example 23-A

10g of ( 2 S , 4 S ) - [ 1- (p-nitrobenzyloxycarbonyl ) -2- { ( 2 - hydroxymethylcarbamoyl ) ethy lmercaptomethyl }-4-acety lthiopyr- rolidine waε diεsolved in 30ml of anhydrous ethyl acetate , and 5m! of trichloroiεocyanate was added dropwise to the resulting solution whi le stirring with ice-cool ing under nitrogen atmosphere . The mixture was continuously stirred for 3 hours under ice-cooling . Then , the reaction solution waε diluted with 50ml of ethyl acetate , waεhed with aqueous εodiu bicarbonate εo lut ion , diεti l led water and sa l i ne ,

respectively, and then dried. The residue was εubjected t column chromatography eluting with ethyl acetate:n hexane(4:l) to obtain the desired product (2S, 4S) -1- (p nitrobenzyloxycarbonyl) -2-{ (2-trichloroacetylaminocarbonyl- oxymethylcarbamoly) ethymercaptomethyl}-4-acetylthiopyrroli- dine in the yield of 52%.

IRfNeatJcm ^"1 : 1755-1750, 1725, 1680, 1600, 1400, 1335 NMR(CDCl ₃ ) S : 1.70-1.95 (3H,m) , 2.45-2.85 (lH,m) , 2.90- 3.15(2H,m), 3.25-3.50(2H,m) , 5.24-5.27

(2H,ε), 7.55-8.27(4H,d,J=8Hz)

Reference Example 23-B

800mg of the compound prepared in Reference Example 23- A waε diεsolved in 5ml of methanol, and 3ml of lN-εodium hydroxide solution waε added dropwiεe to the resulting solution while stirring under ice-cooling. The reaction mixture was stirred for 20 minutes under nitrogen atmos¬ phere, adjusted to the neutral pH value with ^• IN hydrochloric acid solution, and then concentrated under reduced preεsure. The reεidue waε disεolved in ethyl acetate, waεhed with εaline and diεtilled water, respectively, dried and then concentrated under reduced preεsure to obtain an oily resi¬ due. The resulting oily residue waε εubjected to column chromatography eluting with ethyl acetate:n-hexane (5:1) to obtain the desired product (2S,4S)-[1-(p-nitrobenzyloxycar¬ bonyl)-2-{ (2-aminocarbonyloxymethylcarbamoyl)ethylmercapto- methyl}-4-mercaptopyrrolidine in the yield of 65%.

IR(Nujol)cm ^"1 : 1715, 1603, 1512, 1398, 1360 NMR(CDC1 ₃ ) 8 : 1.65-1.95(3H,m) , 2.45-2.95(2H,m) ,

3.01-3.20(3H,m) , 3.25-3.50(2H,m) , 3.84-4.30(2H,m) , 5.24(2H,s), 7.55-8.27

(4H,d,J=8Hz)

Reference Example 24-A

To 21ml of anhydrous N,N-dimethyIformamide were adde 2g of (2S,4R)-1-(p-nitrobenzyloxycarbonyl) -2-{ (2-iodomethyl- carbamoyl)ethylmercaptomethyl}-4-acetylthiopyrrolidine an 1.2g of potasεium phthalimide. The mixture waε εtirred for 6 hourε at 90 to 95°C. The reaction solution was diluted with 50ml of diεtilled water and then extracted three timeε with 50ml of ethyl acetate in each time. The organic layerε were combined, washed with saline and distilled water, respectively, dried and then concentrated under reduced pressure. The residue was subjected to column chromatogra¬ phy eluting with ethyl acetate:n-hexane (2:1) to obtain the desired product (2S,4R)-2-(p-nitrobenzyloxycarbonyl)-2-{ (2- phthalimidoyImethylcarbamoyl)ethylmercaptomethyl}-4-acetyl- thiopyrrolidine in the yield of 72%.

IR(Neat)cm ^-1 : 1775, 1720, 1605, 1522, 1346, 1275 NMR(CDC1 ₃ ) £ : 2.28(3H,s), 2.77(3H,ε), 5.18(2H,ε),

7.46(2H,d,J=8Hz) , 8.16(2H,d,J=8Hz)

Reference Example 24-B

To 20ml of anhydrouε ethanol were added 2.5g of the compound prepared in Reference Example 24-A and 2.3g of hydrazine hydrate. The reaction mixture waε stirred for about one hour under reflux and then cooled to room tempera- ture. The mixture was filtered to remove the insoluble solid materials and then the filtrate was concentrated. The reεidue waε dissolved in 30ml of anhydrous tetrahydrofuran, and 2. Ig of trichloroacetylisocyanate was added to the resulting solution while stirring under ice-cooling. The mixture was stirred for 3 hours and then evaporated under reduced pressure to remove the reaction solvent. The reεi¬ due was subjected to column chromatography eluting with ethyl acetate:n-hexan (3:1) to obtain the desired product

(2S, 4R) -1-(p-nitrobenzyloxycarbonyl) -2-{ (2-trichloroacetyla minocarbonylaminomethylcarbamoyl) ethylmercaptomethyl}-4-ace tylthiopyrrolidine in the yield of 58%.

IR(Neat)cm ^_1 : 1710, 1600, 1517, 1440, 1270

Reference Example 24-C

860mg of the compound prepared in Reference Example 24 B waε dissolved in 10ml of anhydrous methanol, and 1.48ml o lN-sodiu hydroxide εolution waε added dropwiεe to the r εulting εolution while εtirring under ice-cooling. Th reaction mixture waε εtirred for 2 hourε at room tempera ture, neutralized with 1.5ml of IN-hydrochloric acid εolu tion, and then concentrated under reduced presεure. Th reεidue was diluted with 60ml of ethyl acetate, washed wit distilled water and then concentrated again under reduce pressure. The residue was subjected to column chromatograph eluting with ethyl acetate: n-hexane (1:1) to obtain th desired product (2S,4R) -1-(p-nitrobenzyloxycarbonyl) -2—{ (2 aminocarbonylaminomethylcarbamoyl) ethylmercaptomethyl}-4-me rcaptopyrrolidine in the yield of 55%.

IR(Neat)cm ^_1 : 1710, 1665, 1590, 1510, 1425

NMR(CDC1 ₃ ) : 1.75-1.95(3H,m) , 2.45-2.8 (lH,m) ,

2.90-3.15(2H,m) , 3.21(2H,ε), 3.25-3.50 (2H,m), 3.84-4.30(2H,m) , 5.24(2H,m), 7.55-8.27(4H,d,J=8Hz)

Reference Example 25-A

To 25ml of anhydrouε ethanol waε added 1.05g of th compound prepared in Reference Example 24-A. Then, 15ml o saturated ammonia-methanol εolution in anhydrouε methano waε added thereto. The reaction mixture waε stirred for hours under reflux and concentrated under reduced presεure.

The residue was diluted with ethyl acetate, washed with saline and distilled water, reεpectively, and then concen¬ trated under reduced preεεure. The residue was subjected to column chromatography eluting with ethyl acetate:n-hexane (3:1) to obtain the desired product (2S,4R)-1-(p-nitrobenzy¬ loxycarbonyl)-2-{ (2-aminomethylcarbamoyl)ethylmercapto eth- yl}-4-acetylmercaptopyrrolidine in the yield of 82%.

IR(Neat)cm~ ¹ : 1715, 1603, 1512, 1400-1390 NMR(CDC1 ₃ ) 8 : 2.28-2.68 (lH,m) , 3.03-3.70(8H,m) , 3.80-

4.24(2H,m), 5.16(2H,s), 7.49-8.17 (4H,d,J=8Hz)

Reference Example 25-B

To 15ml of anhydrous methylene chloride was added 610mg of the compound prepared in Reference Example 25-A. While εtirring under ice-cooling, 500mg of p-nitrobenzyloxy carbonylchloroformate and 400mg of triethylamine were added to the mixture. The reaction mixture was εtirred for 2 hours and then diluted with 30ml of methylene chloride, washed with IN-hydrochloric acid solution, 10% sodium car¬ bonate solution and distilled water, respectively, dried and then concentrated under reduced pressure. The residue was subjected to column chromatography eluting with ethyl ace¬ tate : n-hexane (4:1) to obtain the desired product (2S, 4R) -1- (p-nitrobenzyloxycarbonyl) -2-{ (2-nitrobenzyloxycarbo- nylaminomethylcarbamoyl) ethylmercaptomethyl}-4-acetylthio- pyrrolidine in the yield of 75%.

IR (Neat) cm ^""1 : 1760, 1715, 1690, 1550, 1480 NMR(CDC1 ₃ ) ' 2.40-3.10(5H,m) , 3.15-3.60 (2H,m) , 5.15-

5.35(4H,m), 7.35-7.70(7H,m) , 7.75-8.15 (2H,m) , .8.22(4H,br.J=8Hz)

Reference Example 25-C

710mg of the compound prepared in Reference Example 25- B waε disεolved in 10ml of anhydrous methanol and thiε reaction mixture waε treated in the εame manner aε that of Reference Example 24-C to obtain the desired product (2S, 4R)-1-(p-nitrobenzyloxycarbonyl)-2-{ (4-nitrobenzyloxycarbon- ylaminomethylcarbamoyl)ethylmercaptomethyl}-4-mercaptopyrro ¬ lidine in the yield of 55%.

IR(Neat)cm~ ¹ : 1710-1700, 1610, 1530-1520, 1350

NMR(CDC1 ₃ ) δ : 1.60-2.00(2H,m) , 2.30-3.65(8H,m) , 3.80-

4.35(2H,m), 5.20(4H,S), 7.50-7.55 (4H,d,J=8Hz) , 8.21(4H,d,J=8Hz)

Reference Example 26-A

1.3g of (2S, 4R) -1-(p-nitrobenzyloxycarbonyl) -2-{ (2- hydroxymethylcarbamoyl)ethylmercaptomethyl}-4-acetylthiopyr- rolidine was added to 30ml of anhydrous methylene chloride. To this reaction mixture were added 2ml of a solution of diazomethane disεolved in ethyl ether and a catalytic amount of boron trifluoride-etherate complex while stirring under ice-cooling, and then the whole mixture waε stirred for 30 minutes. The reaction solution was washed with saline and filtered to remove the" insoluble solid materials. The filtrate was washed with aqueous sodium bicarbonate solu¬ tion, saline and distilled water, respectively. The organic layer waε εeparated and concentrated under reduced preεεure. The residue was subjected to column chromatography eluting with ethyl acetate:n-hexane (10:1) to obtain the desired product (2S,4R)-l-[ (p-nitrobenzyloxycarbonyl) -2-{ (methoxy¬ methylcarbamoyl)ethylmercaptomethyl}-4-acetylmercaptopyrrol- idine in the yield of 62%.

IR(Neat)cm~ ¹ : 1710, 1690, 1685, 1520, 1390 NMR(CDC1 ₃ ) δ = 2.46-2.95(2H,m) , 3.15(3H,s), 3.85-4.15

(3H,m), 5.16(2H,ε), 7.49-8.17(4H,br.J=8Hz)

Reference Example 26-B

910mg of the compound prepared in Reference Example 26- A was added to 15ml of anhydrous methanol, and the reaction mixture was treated in the same manner aε that of Reference Example 24-C to obtain the deεired product (2S, 4R) -1- (p- nitrobenzyloxycarbonyl) -2-{ (methoxymethylcarbamoyl)ethylmer¬ captomethyl}-4-mercaptopyrrolidine in the yield of 50%.

IR(Neat)cm ^_1 : 1710-1700, 1690-1680, 1550-1540, 1390 NMR(CDC1 ₃ ) : 1.60-2.05(2H,m) , 2.33-3.10 (2H,m) , 3.15

(3H,ε) , 3.30-3.80(5H,m) , 3.85-4.33 (2H,m) , 5.24 (2H,ε) , 7.56-8.26 (2H,d,J=8Hz)