DD228258A5 - PROCESS FOR THE PREPARATION OF CARBAPENEM DERIVATIVES - Google Patents

Abstract

A process for the preparation of carbapenem antibiotics having in the 2-position a radical of the general formula SA-R14, wherein A represents a cyclopentylene, cyclohexylene or C2-6-alkylene radical, optionally substituted by one or more C1-4-alkyl groups and R14 represents an aromatic or non-aromatic heterocycle containing a quaternized nitrogen atom attached to A via a quaternary nitrogen atom.

Description

Field of application of the invention

The invention relates to a process for the preparation of carbapenem derivatives. The carbapenem derivatives prepared according to the invention have a substituent of the formula in 2-position

-SAR ¹⁴

wherein A is a cyclopentylene, cyclohexylene or ^ 2 ^ β- alkylene radical, which is optionally substituted by one or more C,, alkyl groups, and Λ U

R is an aromatic or non-aromatic heterocycle containing a quaternized nitrogen atom attached to A via a quaternary nitrogen atom.

Characteristic of the known technical solutions

The carbapenem derivatives prepared by the process of this invention are described in U.S. Patent Application Serial Nos. 366,910, 471,379, and 389,652 (inventors: Choung U. Kim and Peter F, Misco, Jr.). The entire disclosure of each of these applications is hereby incorporated by reference.

US Application 366,910 and its continuation application, Serial No. 471,379 corresponding to DE-OS 3,312,533, describe the preparation of carbapenem antibiotics of the formula

COOR ²

IA

worm

R is a hydrogen atom and R is a hydrogen atom or represents the following substituted and unsubstituted radicals:

an alkyl, alkenyl and alkynyl group of up to 10 carbon atoms; a cycloalkyl and cycloalkylalkyl group having 3 Ms 6 carbon atoms in the cycloalkyl ring and 1 to 6 carbon atoms in the alkyl moieties; a phenyl group; an aralkyl, aralkenyl and aralkinyl group wherein the aryl moiety is a phenyl group and the aliphatic moiety has from 1 to 6 carbon atoms; a heteroaryl, heteroaralkyl, heterocyclyl and heteroeyelylalkyl group, wherein the heteroatom or heteroatoms in the above-mentioned heterocyclic units are selected from 1 to 4 oxygen, nitrogen and sulfur atoms and the alkyl moieties linked to the heterocyclic moieties have 1 to 6 carbon atoms ;

where the substituent or the substituents of the abovementioned radicals is one of the following groups:

a C 1-6 alkyl group, which is optionally substituted by amino, halogen, hydroxy or carboxyl,

a halogen atom,

0 0 NR ³

-OR · ⁵ , -OCNR-TT, -CNR ⁵ R -NR ⁵ R, - < ⁷ ,

^X NR ³ R ⁴ 00

-SO ₂ NR ⁵ R, -NHCNR ⁵ R ^, R-XNR-, -CO ₂ R- ', = 0,

0 0 0

η * * η q η q -χ.

-OCR ⁵ , -SR ⁵ , -SR *, -SR 1, -CN, -N _x , -OSO _x R- *,

-OSO ₅ R ³ , -NR ³ SO ⁴ R ⁴ , -NR ³ C = NR ⁴ , -NR ³ CO ₉ R ⁴ or

r3

-NO ₂ , wherein in the abovementioned substituents the groups

• x. ll

R 1 and R 2 independently represent a hydrogen atom, an alkyl, alkenyl and alkynyl group having 1 to 10 carbon atoms, a cycloalkyl, cycloalkylalkyl and alkylcycloalkyl group having 3 to 6 carbon atoms in the cycloalkyl ring and 1 to 6 Carbon atoms in the alkyl moieties; a phenyl group; an aralkyl, aralkenyl and aralkinyl group wherein the aryl moiety is a phenyl group and the aliphatic moiety has 1 to 6 carbon atoms; or a heteroaryl, heteroaralkyl, heterocyclyl and heterocyclylalkyl group, wherein the heteroatom or heteroatoms in the above-mentioned heterocyclic units are selected from 1 to 4 oxygen, nitrogen or sulfur atoms and the alkyl moieties attached to said heterocyclic moieties have 1 to 6 carbon atoms, or

3 4

R and R together with the nitrogen atom, on

which is bonded to at least one of these radicals, form a 5- or 6-membered, nitrogen-containing, heterocyclic ring;

q *

R ^ is defined as the group R- ^, but none

May mean hydrogen atom; or in which

R and R together represent a Cp ^ Q-alkylidene or hydroxy substituted Cp * Q-alkylidene group;

A is a cyclopentylene, cyclohexylene or a

C ₂ _6 alkylene group, which is optionally substituted by one or more C ₁ _ ^ - alkyl groups;

R is a hydrogen atom, an anionic charge

or a common, easily removable carboxyl-protecting

with the proviso that if R represents a hydrogen atom or a protecting group, a counter anion is also present, and

the rest of the formula

- N

represents a substituted or unsubstituted mono-, bi- or polycyclic aromatic heterocyclic radical containing at least one nitrogen atom in the ring and attached to A via a ring nitrogen atom, thereby forming a quaternary ammonium group;

and their pharmaceutically acceptable acid addition salts. These compounds are prepared by a method given in the following reaction scheme:

  in

IV

COOR

COOR

S -AOH

COOR

2 ¹ diphenylchlorophosphate

HS-A-OH

A = alkylene or Cc-C / -cycloalkylene

methanesulfonyl

SA-OSO ₂ CH ₃

 Ν

COOR

VI

S-A-I

COOR

II

= Counter anion)

COOR

I ¹ A

COOR

IA

US application 389,652 and continuation application 499,690 describe the preparation of carbapenem antibiotics of general formula IB

10 15 20 25

COOR ²

IB

wherein R, R

L Q

R and R are as previously indicated

owning;

R is selected from the following substituted and unsubstituted groups: alkyl, alkenyl and alkynyl of 1 to 10 carbon atoms; Cycloalkyl and cycloalkylalkyl. Having from 3 to 6 carbon atoms in the cycloalkyl ring and from 1 to 6 carbon atoms in the alkyl moieties; phenyl; Aralkyl, aralkenyl and aralkinyl, wherein the aryl moiety is a phenyl radical and the aliphatic moiety has from 1 to 6 carbon atoms; Heteroaryl, heteroaralkyl, heterocyclyl and heterocyclylalkyl wherein the heteroatom or heteroatom in the above-mentioned heterocyclic moieties is selected from 1 to 4 oxygen, nitrogen or sulfur atoms and the alkyl moieties attached to said heterocyclic moieties have 1 to 6 carbon atoms;

wherein the abovementioned R radicals are, if desired, substituted by 1 to 3 substituents which are selected independently of one another from

a C, g alkyl radical which is optionally substituted by amino, fluoro, chloro, carboxyl, hydroxy or carbamoyl;

Fluorine, chlorine or bromine;

-OR ³ J -OCO ₂ R ³ ; -OCOR ³ ; -OCONR ³ R ⁴ ; -OSO ₂ R ³ ;

oxo; -NR ³ R ⁴ ; R ³ CONR ⁴ -; -NR ³ CO ₂ R ⁴ ;

0 -NR ³ CONR ³ R ⁴ ; -NR ³ SO ₂ R ⁴ ; -SR ³ ; -SR ⁹ ;

-SR ⁹ ; -SO ₃ R ³ ; -CO ₂ R ³ ; -CONR ³ R ⁴ ; -CN; or

a phenyl radical which is optionally substituted by: 1-3-fluoro, chloro, bromo, C 1-6 alkyl, -OR ³ , -NR ³ R ⁴ , -SO, R ³ , -CO ₉ R ³ or -CONR ³ R ⁴ , wherein R ³ , R and R in these substituents R angege the above

possess meanings; or in which R can denote a C 1-6 -alkylene group which is in the ring:

or wherein R is a divalent phenylene or

θ s ~ \

connected so as to form a bridged, polycyclic group;

A represents a cyclopentylene or cyclohexylene radical or a C ₂ _g alkylene group, which is optionally substituted by one or more C ₁ alkyl groups;

R is a hydrogen atom, an anionic charge

or a common, easily removable carboxyl-protecting

with the proviso that when R represents a hydrogen atom or a protecting group, a counterion is also present; and the rest of the formula

a substituted or unsubstituted, mono-, bi- or polycyclic, non-aromatic heterocyclic radical containing at least one nitrogen atom in the ring and attached to A via a ring nitrogen atom, thereby forming a quaternary ammonium group;

and the pharmaceutically acceptable salts thereof. This process is shown in the following reaction scheme:

, diphenyl

COOR

III

IV ..

0 Il OP (OC ₆ H ₅ ):

COOR

HS-A-OH

A = alkylene or C ₅ -Cg -cycloalkylene

S-A-OH

Methanesulfonyl chloride

COÖR

SA-OSO ₂ CH ₃

COOR

VI

,8th

-N

II

I ¹ B

IB

10

S-A-I

COOR

S-A-

COOR

if necessary, blocking .

In this known method is based on the compound of formula III and sets them in an inert organic solvent with diphenylchlorophosphate in the presence of a base to the intermediate IV. The intermediate compound IV is then with a

Mercaptan reagent of the formula HS-A-OH in an inert organic solvent and in the presence of a base to the intermediate V. Intermediate compound V is then acylated with methanesulfonyl chloride in an inert organic solvent and in the presence of a base to give intermediate VI, which is reacted with an iodide ion source in an inert organic solvent to give intermediate II. This intermediate II is reacted with the desired amine in an inert organic solvent and in the presence of silver ions to give the quaternized product I ¹ A or I ¹ B which can then be deblocked to give the corresponding deblocked carbapenem compound of the formula IA or IB receives.

The method described above has several disadvantages.

So this process goes through different stages, whereby it would be beneficial if you could reduce the number of stages. The yield of the overall reaction is also quite low. In addition, the quaternization is carried out with the entire carbapenem compound.

Object of the invention

The aim of the present invention is therefore to provide a novel process for the preparation of compounds IA or IB which (1) has fewer reaction steps, (2) leads to higher yields, (3) first allows the formation of the quaternized amine, which is later (4) allows easier formation of quaternary amine products with a variety of amines, eg, sterically hindered or low valent amines.

According to the invention, a novel process for the preparation of carbapenem derivatives of the general formula I is provided:

, 8 H

S-A-R

COOR '

14

wherein

R is a hydrogen atom and, 1

R 'represents a hydrogen atom or represents the following substituted and unsubstituted radicals:

an alkyl, alkenyl and alkynyl group of up to 10 carbon atoms; a cycloalkyl and cycloalkylalkyl group having 3 to 6 carbon atoms in the cycloalkyl ring and 1 to 6 carbon atoms in the alkyl moieties; a phenyl group; an aralkyl, aralkenyl and ar alkynyl group wherein the aryl moiety is a phenyl group and the aliphatic moiety has from 1 to 6 carbon atoms; a heteroaryl, heteroaralkyl, heterocyclyl and heterocyclylalkyl group wherein the heteroatom or heteroatoms in the above-mentioned heterocyclic moieties are selected from 1 to 4 oxygen, nitrogen and sulfur atoms and the alkyl moieties attached to the heterocyclic moieties have 1 to 6 carbon atoms ;

the. Substituent or substituents of the abovementioned radicals mean one of the following groups:

a C, g-alkyl group, which is optionally substituted by amino, halogen, hydroxy or carboxyl,

a halogen atom,

0 0 -OR ³ , -OCNR ³ R ⁴ , -CNR ³ R ⁴ , -NR ³ R ⁴ , - (^

0 0

-SO ₂ NR ⁵ R, -NHCNR ⁵ R, R ⁵ CNR-, -CO ₂ R ⁵ , = 0, 0 0 0

-OCR ³ , -SR ³ , -SR ⁹ , -SR ⁹ , -CN, -N ,, -OSO ^ R ³ ,

πJ D

-OSO ₂ R ³ , -NR ³ SO ₂ R ⁴ , -NR ³ C = NR ⁴ , -NR ³ CO ₂ R ⁴ or

-NO ₂ , wherein in the abovementioned substituents the groups

• ζ Λ

R 1 and R 4 independently represent a hydrogen atom, an alkyl, alkenyl and alkynyl group of 1 to 10 carbon atoms, a cycloalkyl, cycloalkylalkyl and alkylcycloalkyl group of 3 to 6 carbon atoms in the cycloalkyl ring and 1 to 6 carbon atoms in the alkyl moieties; a phenyl group; an aralkyl, aralkenyl and aralkinyl group wherein the aryl moiety is a phenyl group and the aliphatic portion has 1 to 6 carbon atoms, or a heteroaryl, heteroaralkyl, heterocyclyl and heterocyclylalkyl group, wherein the heteroatom or heteroatoms in the above-mentioned heterocyclic units are selected from 1 to 4 oxygen, nitrogen or sulfur atoms and the alkyl moieties attached to said heterocyclic moieties have 1 to 6 carbon atoms, are, or

IA

3 4

R and R together with the nitrogen atom, on

the at least one of these radicals is bonded, a 5- or 6-membered, nitrogen-containing, hetero-:

form cyclic ring;

ο 3

R is defined as the group R, but no

May mean hydrogen atom; or in which

R and R together represent a Cp ^ Q-alkylidene or hydroxy-substituted Cp * Q-alkylidene radical:

A is a cyclopentylene, cyclohexylene or a

C ₂ _g-alkylene group, which is optionally substituted by one or more C ^^ - alkyl groups;

R is a hydrogen atom, an anionic charge

or a common, easily removable carboxyl-protecting

with the proviso that, if R represents a hydrogen atom or a protecting group, also

20 · there is a counteranion, and

14

R is a quat te mi si er th, nitrogen-containing, aromatic or non-aromatic heterocycle, which is bound via a ring nitrogen atom to A, whereby a quaternary Ammo nium group is formed,

or a pharmaceutically acceptable salt thereof, which is characterized in that

an intermediate of general formula IV 30

O- ^X C00R ²

IV

In which R and R have the meanings given above

2 '

R, represents a common, easily removable carboxyl-protecting group and L represents a common leaving group, with a thiol compound of general formula VII

HS-A-R VII

14

, 14

wherein A and R have the meanings given above and X ° represents a counter anion, in an inert solvent and in the presence of a base to a Carbap enemverbindung the general formula I ^1st

S-A-R

wherein R ¹ , R ⁸ , R

2'14 ft

, A, R and Χ ^σ are those given above

Have 25 meanings,

converts and desired! "alls the carboxyl-protecting group

2 '

R to obtain the corresponding deblocked compound of general formula I or a pharmaceutically acceptable salt thereof. 30

The invention also relates to intermediate compounds of general formula VII and to processes for preparing these intermediates.

10

The carbapenem compounds of general formula I are effective antibacterial agents or intermediates useful for preparing such agents.

15

30

The compounds of general formula I contain the carbapenem nucleus

and thus may be referred to as i-carba-p-phen-S-carboxylic acid derivatives. However, the compounds may alternatively be considered to have the following basic structure

and are referred to as 7-0xo-1-azabicyclo [3.2.0] hept-2-ene-2-carboxylic acid derivatives. According to the invention compounds are included, wherein the hydrogen atoms in the 5- and 6-position can be present in both the cis and in the trans position, the preferred compounds having 5R, 6S (trans) configuration, as is the case with thienamycin is.

35

The compounds of the general formula I may be unsubstituted in the 6-position or substituted by the substituent groups described above for the other carbapenem derivatives. In particular, means

8 1

R represents a hydrogen atom and R may represent a hydrogen atom or a substituent other than hydrogen, and for example in European Patent Application 38,869 (see the definition of

* 8 R /) is disclosed. Alternatively, R and R may together form a Cp. ₀ alkylidene radical or a substituted (eg by hydroxy) C _2-1 Q-alkylidene radical.

1 Q

The definitions for R and R are described below:

(a) Aliphatic alkyl, alkenyl and alkynyl groups are straight-chain or branched groups having 1 to 10 carbon atoms, preferably 1 to 6 and especially preferably 1 to 4 carbon atoms. If these groups are part of another substituent, for example in a cycloalkyl radical or a heteroaralkyl or aralkenyl radical, then the alkyl, alkenyl and alkynyl group preferably contains 1 to 6 and more preferably 1 to 4 carbon atoms.

(b) By "heteroaryl" is meant mono-, bi- and polycyclic aromatic heterocyclic groups containing 1 to 4 O, N or S atoms. Preferred are 5- or 6-membered heterocyclic rings such as thienyl, furyl, thiadiazolyl, oxadiazolyl, triazolyl, isothiazolyl, thiazolyl, imidazolyl, isoxazolyl, tetrazolyl, oxazolyl, pyridyl, pyrazinyl, pyrimidinyl, pyridazinyl, pyrrolyl, pyrazolyl, etc.

(C) By "heterocyclyl" is meant mono-, bi- and poly-oleic, saturated or unsaturated, non-aromatic, heterocyclic groups containing 1 to 4 O, N or S atoms. Preference is given to 5- or 6-membered heterocyclic rings, such as morpholinyl, piperazinyl, piperidyl, pyrazolinyl, pyrazolidinyl, imidazolinyl I, imidazolidinyl, pyrrolinyl, pyrrolidinyl, etc. 10

(d) "halogen" refers to chlorine, bromine, fluorine and iodine atoms and preferably chlorine or bromine atoms.

The term "common, readily removable, carboxyl-protecting group" refers to a known ester group which has been used to block a carboxyl group during the chemical reactions described below, and which can, if desired, be removed by methods which do not destroy the remainder of the molecule. Such methods include, for example, chemical or enzymatic hydrolysis, treatment under mild conditions with chemical reducing agents, irradiation with UV light, and catalytic hydrogenation. Examples of such ester-forming protecting groups are benzhydryl, p-nitrobenzyl, 2-naphthylmethyl, allyl, benzyl, trichloroethyl, silyl, e.g. Trimethylsilyl, phenacyl, p -methoxybenzyl, acetonyl, ο-nitrobenzyl, 4-pyridylmethyl and Cj_g-alkyl, e.g. Methyl, ethyl or t-butyl. Protecting groups are also those which are hydrolyzed under physiological conditions. These include, for example, pivaloyloxymethyl, acetoxymethyl, phthalidyl, indanyl and methoxymethyl. Particularly advantageous carboxyl-protecting groups are the p-nitrobenzyl group, which are easily removed by catalytic hydrogenolysis

can, and the allyl group, which can be removed by Pd (P0 ^) catalyzed reaction. 5

The pharmaceutically acceptable salts include the non-toxic acid addition salts, e.g. Salts with mineral acid, such as hydrochloric acid, hydrobromic acid, hydroiodic acid, phosphoric acid, sulfuric acid, etc., and salts with organic acids, such as maleic, acetic, citric, succinic, benzoic, tartaric, fumaric, almond, Ascorbic, lactic, gluconic and malic acid. Compounds of general formula I in the form of the acid addition salts can be described as

SAR ¹⁴

ρ, R = H or a protecting group

θ Α

wherein X represents the acid anion. The counter anion X can be selected so that pharmaceutically acceptable salts are obtained for therapeutic purposes. In the case of the intermediate compounds of the formula I, however, X can also be a toxic anion. In such a case, the ion may subsequently be removed or replaced with a pharmaceutically acceptable anion to yield the active end product for therapeutic use.

1 14

If acidic or basic groups are present in the group R or on the quaternized R residue, suitable base or acid salts of these functional groups are also included according to the invention, e.g. Acid addition salts of a basic group and metal salts (e.g., sodium, potassium, calcium and aluminum), the ammonium

salt and salts with nontoxic amines (for example trialkylamine, procaine, dibenzylamine, 1-ephenamine, N-benzyl-ßphenethylamin, Ν, Ν ¹ - dibenzylethylenediamine, etc.) in · an acidic group.

Compounds of the general formula I in which R represents a hydrogen atom, an anionic charge or a physiologically hydrolyzable ester group, and the pharmaceutically acceptable salts thereof are useful antibacterial agents. The other compounds of the general formula I are valuable intermediates which can be found in the abovementioned biologically active compounds can be converted.

Preferred compounds according to the invention are those compounds of the general formula I in which R is a hydrogen atom and R is a hydrogen atom, 20

OH is CH-, 'C- or CH ₃ CH-.

CH, ^

-j 3

5 In this subclass are the preferred compounds

OH

1 '

those in which R is CH 1 CH-. Particularly preferred compounds have the absolute configuration 5R, 6S, 8R.

Preference is furthermore given to those compounds of the general formula I in which R and R together form an alkylidene radical of the general formula

HIGH ₂

C =

form.

The alkylene or cycloalkylene radical A in the compounds of the general formula I can be a cyclopentylene radical

a cyclohexylene radical

or a C ₂ _g alkylene radical, which is optionally substituted by one or more C ₁ ^ alkyl substituents. Preferred substituents A are cyclopentylene, cyclohexylene or alkylene of the formula

R ¹⁰ R ¹²

-C-

R ¹¹ R ¹³

wherein R ¹⁰ , R ¹¹ , R ¹² and R ¹ x ⁵ each independently represent a hydrogen atom or a C ₁ λ -Alky lgruppe. A preferred embodiment comprises such

Compounds of general formula I, wherein the substituent A is -CH CH _{0 9} -, -CHCH ₀ -, - /> CH- or -CH ₀

ptl > ' _nV s

^ n₃ ' OH₃

stands.

In some compounds of general formula I, when substituent A is a cycloalkylene or branched alkylene radical, one or more additional asymmetric carbon atoms are present resulting in the formation of diastereoisomers. According to the invention, the mixtures of these diastereoisomers and also the individual, purified diastereoisomers are claimed.

The quaternized substituent R is an optionally substituted, mono-, bi- or polycyclic, aromatic or non-aromatic, heteroalkyl radical containing at least one nitrogen atom in the ring and attached atom A via a ring nitrogen, thereby forming a quaternary ammonium group ,

A preferred class of substituents R are those of the general formula

-O

This formula represents a substituted or unsubstituted, mono-, bi- or polycyclic heteroaryl radical which contains at least one nitrogen atom in the ring and is bonded via a ring nitrogen atom to a carbon atom of the substituent A, resulting in a quaternary ammonium group. The heteroaryl radical may, if desired, be substituted by the following substituents:

ated be: C ₁ ^ -AIkVIj C ^^ - alkyl substituted by hydroxy, amino, carboxy or halogen; C, C 1 -cycloalkyl; _1- C ^ -alkoxy; C ₁ _ ^ - alkylthio; amino; C _{1 to} C alkylamino; Di- (C ₁ _ ^ - alkyl) amino; Halogen; Cj ^ alkanoylamino; C ₁ _ ^ - alkanoyloxy; carboxy;

O-C-OC _{1 -Zf} -alkyi; hydroxy; amidino; Guanidino, trifluoromethyl; phenyl; Phenyl substituted with one, two, or three amino, halo, hydroxy, trifluoromethyl, C _{1 -Zj>} alkyl, or C 1-10 alkoxy groups; Heteroaryl and heteroaralkyl, wherein the heteroatom or heteroatoms in the above-mentioned heterocyclic units are selected from 1 to 4 O, N or S atoms and the alkyl group connected to the heteroaralkyl group has 1 to 6 carbon atoms.

The heteroaryl group attached to the substituent A is preferably a 5- or 6-membered aromatic heterocyclic group containing a quaternized nitrogen atom bonded directly to a carbon atom of the alkylene or cycloalkylene group, and optionally one or more additional heteroatoms selected Although generally each heteroaryl radical attached to A via a quaternized nitrogen atom results in biologically active carbapenem derivatives, a preferred embodiment of the invention comprises compounds wherein the radical of the general formula

one of the radicals listed below: 35

to

(A)

wherein R, R and R are independently

"Hydrogen; C 1-6 alkyl; C 1-10 alkyl substituted by hydroxy, amino, carboxy or halo; C 1-6 cycloalkyl; C 1-10 alkoxy; C 1-10 -alkylthio; amino; C 1-8- Alkylamino; di- (C ₁ -C 4 -alkyl) '- amino; halogen; C 1 -C 4 -alkanoylamirio; C ^ _ ^ -

O

alkanoyloxy; carboxy; -C-OC ₁ _ ^ - alkyl; hydroxy; amidino;

guanidino; trifluoromethyl; phenyl; Phenyl substituted by one, two or three amino, halo, hydroxy, trifluoromethyl, C .._ ^ - alkyl or C ₁ »alkoxy groups; and heteroaryl and heteroaralkyl wherein the heteroatom or heteroatoms in the above-mentioned heterocyclic units are selected from 1 to 4 oxygen, nitrogen or sulfur atoms and the alkyl moiety linked to said heteroaralkyl moiety has from 1 to 6 carbon atoms; or wherein two of R, R or R together form a fused saturated carbocyclic ring, a fused aromatic carbocyclic ring, a fused saturated heterocyclic ring or a fused heteroaromatic ring;

30

or

35

10 15 20 25

wherein these radicals are optionally substituted on a carbon atom by one or more substituents independently selected from the following radicals: C 1 -C 4 alkyl; C 1-4 alkyl substituted by hydroxy, amino, carboxy or halogen; C 1-6 cycloalkyl; C ₁ »-alkoxy; C._λ-alkylthio; amino; C ₁ _λ-alkylamino; Di- (C ._, - alkyl) amino; Halogen; C. ^ -Alka-

11

noyloxy; C ₁ λ-alkanoylamino; carboxy; -C-OC ₁ ^ -alkyl; hydroxy; amidino; guanidino; trifluoromethyl; phenyl; Phenyl substituted by one, two or three amino, halo, hydroxy, trifluoromethyl, C ^^ - alkyl or C ₁ a-alkoxy groups; and heteroaryl or heteroaralkyl wherein the heteroatom or heteroatoms in the above-mentioned heterocyclic units are selected from 1 to 4 oxygen, nitrogen or sulfur atoms and the alkyl moiety attached to said heteroaralkyl moiety has from 1 to 6 carbon atoms, or if so substituted, that a fused carbocyclic or heteroethylene ring is formed; . - ...

(C)

30 35

or

wherein these radicals are optionally substituted on a carbon atom by one or more substituents independently selected from the following radicals: C 1 -C 4 alkyl; C 1-8 alkyl substituted by hydroxy, amino, carboxy or halogen; C, _ £ cycloalkyl; C ₁ ^ -alkoxy; C ^^ - alkylthio; amino; C ₁ _r-alkylamino; Di- (C ₁ - alkyl) amino; Halogen; C ₁ r alkanoylamino; C 1-4 alkanoyloxy; carboxy; 0

Il

-C-OC ₁ _ ^ - alkyl; Hydroxy, amidino; guanidino; trifluoromethyl; phenyl; Phenyl substituted by one, two or three amino, halo, hydroxy, trifluoromethyl, C _1- / -alkyl or C ₁ ^ alkoxy groups; and heteroaryl or heteroaralkyl wherein the heteroatom or heteroatoms in the above-mentioned heterocyclic units are selected from 1 to 4 oxygen, nitrogen or sulfur atoms and the alkyl moiety attached to said heteroaralkyl moiety has from 1 to 6 carbon atoms, or if so substituted, that a condensed carbocyclic or heterocyclic ring is formed;

, S *

or

these radicals, if desired, on a carbon. , Substituted by one or more substituents which are independently selected from the following radicals: C ^^ - alkyl; C _{1 to} C alkyl substituted by hydroxy, amino, carboxy or halogen; C, C 1 -cycloalkyl; C ₁ ^ -alkoxy; C _1- Zf-alkyl thio; amino; C ^^ - alkyl amino; Di- (C ₁ -alkyl) -amino, halo, C ₁ -alkanoylamino; C 1-10 alkanoyloxy, carboxy;

-C-OC ₁ _ ^ - alkyl; hydroxy; amidino; guanidino; trifluoromethyl; phenyl; Phenyl substituted by one, two or three amino, halo, hydroxy, trifluoromethyl, C ₁ _ ^ - alkyl or C _1- ^ -AIkOXygruppen; and heteroaryl or heteroaralkyl, wherein the heteroatom or heteroatoms in the above-mentioned heterocyclic units are selected from 1 to 4 oxygen, nitrogen or sulfur atoms and the alkyl moiety linked to said heteroaralkyl moiety has 1 to 6 carbon atoms, or are so desirably substituted in that a condensed carbocyclic or heterocyclic ring is formed;

(E)

or

wherein X is 0, S or NR, wherein R is C _1- ^ -alkyl or phenyl, these radicals being optionally substituted on a carbon atom by one or more substituents which are independently selected from the following radicals: C ₁ _ ^ alkyl; C _1- Zt-alkyl, substituted by hydroxy, amino, carboxy

or halogen; C 1-5 cycloalkyl; Cj ^ alkoxy; C ^^ thio; amino; C. _, - alkylamino; Di (C 1-6 alkyl) amino; Halogen; C ₁ _, alkanoylamino; C ,,, alkanoyloxy; carboxy; 0

Il

-C-OC ₁ _ _{/ f} -alkyl; hydroxy; amidino; Guanidinoj trifluoromethyl; phenyl; Phenyl substituted by one, two or three amino, halo, hydroxy, trifluoromethyl, C ₁ H 4 alkyl or C ₁ H 4 alkoxy groups; and heteroaryl or heteroaralkyl wherein the heteroatom or heteroatoms in the above-mentioned heterocyclic units are selected from 1 to 4 oxygen, nitrogen or sulfur atoms and the alkyl moiety attached to said heteroaralkyl moiety has from 1 to 6 carbon atoms, or if so substituted, that a condensed carbocyclic or heterocyclic ring is formed;

^(f) / 7 λ

^ X X

*

X N-. N or

N-

wherein X is 0, S or NR, wherein R is C ₁ ^ alkyl or phenyl, these radicals being optionally on a carbon atom by one or more substituents which are independently selected from the following radicals: C ₁ ^ - AIlCyI; C ₁ _ _Zf alkyl substituted by hydroxy, 'amino, carboxy or halogen; C ₃ _ ₆ cycloalkyl; C ^ alkoxy; C ₁ -Zf alkyl

thio; amino; C ₁ _ _Zf alkylamino; Di- _(C1 ^ alkyl) amino; Halogen; C ^ _ ^ - alkanoylamino; C ₁ -Zf alkanoyloxy; carboxy; 0

Il

-C-OC ₁ _ ^ - alkyl; hydroxy; amidino; guanidino; trifluoromethyl; phenyl; Phenyl substituted with one, two or three amino, halo, hydroxy, trifluoromethyl, C ₁ l -alkyl or C ₁ / alkoxy groups; and heteroaryl or heteroaralkyl, wherein the heteroatom or heteroatoms in the above-mentioned heterocyclic units are selected from 1 to 4 oxygen, nitrogen or sulfur atoms and the alkyl moiety linked to said heteroaralkyl moiety has from 1 to 6 carbon atoms; and

NNR%

N N-R

or «· '

wherein R is C 1-4 alkyl or phenyl, which radicals are optionally substituted on a carbon atom by one or more substituents independently of one another from the following radicals

are selected: C _1- ^ alkyl; C 1-10 alkyl substituted by hydroxy, amino, carboxy or halogen; CU / cycloalkyl; C ^ _ ^ - alkoxy; C ^^ - alkylthio; amino; C ^ ^ -Alky! Amino; Di- (C _r alkyl.) Amino; C. _ <- alkanoylamino; 0

Il

carboxy; -C-OC ₁ ^ -alkyl; hydroxy; amidino; guanidino; trifluoromethyl; phenyl; Phenyl substituted by one, two or three amino, halo, hydroxy, trifluoromethanesulfonic tiiyl-, C ₁ _4 ~ alkyl or C ^ alkoxy; and heteroaryl or heteroaralkyl, wherein the heteroatom or heteroatoms in the above-mentioned heterocyclic moieties are selected from 1 to 4 oxygen, nitrogen or sulfur atoms and the alkyl moiety attached to said heteroaralkyl moiety has 1 to 6 carbon atoms.

In the above subclass, the preferred compounds are those in which the substituent A is -CH ₂ CH ₂ -, -CHCH ₂ -,

- / S or -CH ₂ CH-, and wherein - ^ CH ₃

1 8 5 (a) R and R together for

CH,

HIGH ₂

CH ₃

C =

stand or

(b) R ^{8 is} hydrogen and R is for

a hydrogen atom, CH ₃ CH ₂ -,

'CH ₃ OH

CH-, ^ C- or CH _x CH- stands. CH ₃ CH ₃

CH ₃ 'CH ₃ OH OH

Particular preference is given to compounds in which R is a

OH

1 'is hydrogen atom and R is CH, CH-, the preferred compounds having the absolute configuration 5R, 6S, 8R.

According to a particularly preferred embodiment of the invention, compounds are obtained in which the rest of the formula ...

-Ό

a remainder of the formula

R ⁶

5 6 7, wherein R, R and R independently represent a hydrogen atom, a C ^ alkyl group, a C ._ ^ - alkoxy group, a C ₁ ^ alkyl group substituted by a hydroxy group, a C ₁ _ ^ Alkylthio group, an amino, carboxy and carbamoyl group. In this subclass, the preferred compounds are

before, the substituent A is -CH ₉ CH ₅ -, -CHCH ₉ -,

- / \ or -CH ₂ CH- and in which either

'CH,

d R ⁸ zi

HIGH

(a) R and R together for

CH,

stand or

R 1

(b) R is a hydrogen atom and R is hydrogen, CH ₃ CH ₂ -,

CH,. CH, OH OH

CH, C or CH ₃ CH- stands.

CH _{3 3}

Especially preferred are compounds wherein R is a

OH 1 1

Is hydrogen and R is CH ₃ CH-. The

preferred compounds have the absolute configuration 5R, 6S, 8R

 15

A most preferred embodiment of the invention involves the preparation of compounds wherein the radical of the general formula

- Ν)

a radical of the general formula

, 6

C C ty

in which R, R and R 'independently represent a hydrogen atom, a C ₁ ^ alkyl, C ^ ^ alkoxy, by a hydroxy-group-substituted C ^^ - alkyl, C ₁ Λ-alkylthio and amino mean. In this subclass, the preferred compounds are those in which the substituent A is -CH ₂ CH ₂ -, -CHCH ₂ -,

- (\ or -CH ₂ CH- and in which either

CH ₃ ·

(a) R and R together for

HIGH ₂

C =

CH,

stand or

O A

(b) R is a hydrogen atom and R is hydrogen, CH ₃ CH ₂ -,

CH, CH, OH OH

CH, C or CH ^ CH- stands. /

₃ CH ₃

Especially preferred are compounds wherein R is for

OH

Is hydrogen and R ^{1 is} CH ₃ CH-. The preferred compounds have the absolute configuration 5R, 6S, 8R.

Another preferred embodiment of the invention comprises the preparation of compounds in the radical of the general formula

-O

a radical of the general formula

SCH,

means.

In this subclass preferred compounds are those in which the substituent A is -CHCH ₂ -, -CH ₂ CH ₂ -,

CH ₃ 35

- / \ or -CHpCH- and in which either

} - ητι

CH ₃

(a) R land R together for

HIGH ₂

stand or

(b) R represents a hydrogen atom and R ^{1 represents}

CH., CH-OH

Hydrogen, CH ₃ CH ₂ -, CH- \ c- or;

'. ^C1 C ^C1 C

OH CH ₃ CH- stands.

In particular: preference is given to compounds in which R is an OH hydrogen atom and R is CH ₃ CH-. The preferred compounds have the absolute configuration 5R, 6S, 8R,

Another preferred embodiment of the invention comprises the preparation of compounds wherein the radical of the general formula

-O

represents a pyridinium radical. In this subclass, preferred compounds are those wherein the substituent

tuent A is -CHCH ₂ -, -CH ₂ CH ₂ -, - ^^ or -CH ₂ CH- is CH ₃ CH ₃

and. in which either

(a) R and R together for

-C =

CH-

stand or

8 1

(b) R is a hydrogen atom and R is for

CH,

CH-

Hydrogen, CH ₃ CH ₂ , ^x CH-

OH ³

15 CH ₃ CH- stands.

OH

C or

Particular preference is given to compounds in which R is a

OH 1 '

means hydrogen and R is CH, CH-. The preferred compounds have the absolute configuration 5R, 6S, 8R.

A most preferred embodiment of the invention involves the preparation of compounds of the general formula

OH H

(R)

-S-A

-O

• COOR '

wherein the radical of the general formula -A-!

has the following meanings:

- N

CH,

CH

(5) -CH ₂ CH ₂ - ^ N

3 CH.

OCH.

(13) -

(2)

(4)

(10)

CHCH. ι * CH-,

R- or S-diastereoisomers

(12)

or (14)

-CH ₉ CH-

CH ₂ OH

(6) -CH ₂ CH ₂ -

SCH.

CH ₂ OH ⁽⁸ *. "" ^CH 2 ^CH 2 ~

SCH.

R. _f R or S, S-diastereoisomers on the two asymmetric carbon atoms of the cyclohexyl group

In which R is a water atom, an anionic charge or a common, easily removable carboxyl

p represents a protecting group, provided that if R

represents a hydrogen atom or a protective group, also a Gegenanion is present, and the pharmaceutically acceptable acid addition salts thereof.

A still preferred class of quaternized

14 R substituents are those of the general formula

R ¹⁵

Wherein R is selected from the following substituted and unsubstituted radicals: alkyl »alkenyl and alkynyl having 1 to 10 carbon atoms; Cycloalkyl and cycloalkylalkyl of 3 to 6 carbon atoms in the cycloalkyl ring and 1 to 6 carbon atoms in the alkyls; phenyl; Aralkyl, aralkenyl and aralkinyl, wherein the aryl moiety is phenyl and the aliphatic moiety has from 1 to 6 carbon atoms; Heteroaryl, heteroaralkyl, heterocyclyl and heteroeyelylalkyl wherein the heteroatom or heteroatoms in the above-mentioned heterocyclic moieties are selected from 1 to 4 oxygen, nitrogen or sulfur atoms and the alkyl moieties attached to said heterocyclic moieties have from 1 to 6 carbon atoms; where the abovementioned radicals R are, if desired, substituted by 1 to 3 substituents, which are selected independently of one another from the following radicals:

C, g-alkyl, optionally substituted by amino, fluoro, chloro, carboxyl, hydroxy or carbamoyl;

Fluorine, chlorine or bromine;

-OR ⁵ } -OCO ₂ R ³ ; -OCOR ³ ; -OCONR ³ R ⁴ ; -OSO ₂ R ³ ;

oxo; -NR ³ R ⁴ ; R ³ CONR ⁴ -; -NR ³ CO ₂ R ⁴ ;

-NR ³ CONR ³ R ⁴ ; -NR ³ SO ₂ R ⁴ ; -SR ³ ; -SR ⁹ ; 0.0 -SR ⁹ ; -SO ₃ R ³ ; -CO ₂ R ³ ; -CONR ³ R ⁴ ; -CN; or

a phenyl radical, if desired substi

is substituted by: 1-3-fluoro, chloro, bromo, -OR ³ , -NR ³ R ⁴ , -SO ₃ R ³ , -CO ₂ R ³ or -CONR ³ R ⁴ , wherein R ³ , R and R in these substituents R have the meanings given above;

or in which

16 R is a divalent phenylene or C ₁ , -alkylene

group means which with the

Ring is connected so that forms a bridged, polycyclic group, and

the rest of the general formula - N

a substituted or unsubstituted, mono-, bi- or polycyclic, non-aromatic, heterocyclic radical which may be fused to another aromatic or non-aromatic ring and which has at least one nitrogen atom in the ring and bound via a ring nitrogen atom to A. is, resulting in a quaternary / ammonium group. The heterocyclic radical can be saturated or unsaturated

(having 1 to 2 double bonds) and may contain up to two further heteroatoms in addition to the quaternary nitrogen atom. Such additional heteroatoms are O, S (0) _mt N, NR ¹⁵ or NR ¹⁷ R ¹⁸ wherein m is 0, 1 or 2, R ^{1 is} hydrogen, optionally substituted C ₁ -alkyl or optionally substituted Phenyl group and R and R ¹⁸ independently represent an optionally substituted C ₁ g alkyl or optionally substituted phenyl group.

In a preferred embodiment, the radical is of the general formula

a non-aromatic, 4- to 7-membered, preferably 5- or 6-membered, N-containing, heterocyclic ring having 0 to 2 double bonds and 0 to 2 additional heteroatoms, which is less than 0, S (0) _m , N, NR ¹⁵ or NR ¹⁷ R ¹⁸ is selected, wherein m is 0, 1 or 2, R ¹⁵ represents a hydrogen atom, a C _1-6 alkyl group which is substituted if desired, by 1 to 2 substituents, where these substituents are independently selected under -OR ³ , -NR ³ R ⁴ , -CO ₂ R ³ , oxo, phenyl, fluorine, chlorine, bromine, -SO ₃ R ³ and -CONR ³ R, or phenyl group which is optionally substituted by 1 to 3 substituents where these substituents are independently selected from C 1-6 -alkyl, -OR ³ , -NR ³ R ⁴ , fluorine, chlorine, bromine, -SO ₃ R ³ , -CO ₂ R ² or -CONR ³ R ⁴ , and R ¹⁷ and R ¹⁸ are each independently a C ₁ _g alkyl group which is substituted if desired, by 1 to 2 substituents, which Substi are independently selected from -OR ³ , -NR ³ R ⁴ , -CO ₉ R ³ ,

Oxo, phenyl, fluorine, chlorine, bromine, -SO ^ R ³ and -CONlAl, or a phenyl group which is optionally substituted by 1 to 3 substituents, these substituents independently of each other

-z. -z h

are selected from C 1-6 alkyl, -OR, -NR 1-4 R, fluoro, chloro, bromo, -SO ^ R ³ , -CO ₂ R ² and -CONR ³ R ⁴ , wherein R ³ and R in these heterocyclic NR ¹ - * - or NR ¹ ^ R ¹⁸ groups have the meanings given above in connection with the substituent R. In such a preferred embodiment, the ring

If desired, be substituted by 1 to 3 of the following substituents:

(a) C, g-alkyl, optionally substituted by T to 2 substituents independently selected from fluoro, chloro, bromo, -OR, -OCOR ³ , -OCONR ³ R ⁴ , oxo, -NR ³ R ⁴ , -NR ³ COR ⁴ , -NR ³ CONR ³ R ⁴ ,

-NR ³ SO ₂ R ⁴ , -SR ³ , -SO ₃ R ³ , -CO ₂ R ³ and -CONR ³ R ⁴ ;

(b) C ₂ _g alkenyl, optionally substituted with 1 to 2 substituents independently selected from fluoro, chloro, bromo, -OR ³ , -OCOR ³ , -OCONR ³ R ⁴ , oxo, -NR ³ R ⁴ , -NR ³ COR ⁴ , -NR ³ CONR ³ R ⁴ ,

-NR ³ SO ₂ R ⁴ , -SR ³ , -SO, R ³ , -CO ₂ R ³ and -CONR ³ R ⁴ ;

(c) C ₂ _g-alkynyl, optionally substituted by 1 to 2 substituents independently selected from fluoro, chloro, bromo, -OR, -OCOR ³ , -OCONR ³ R ⁴ , oxo, -NR ³ R ⁴ , -NR ³ COR ⁴ , -NR ³ CONR ³ R ⁴ ,

-NR ³ SO ₂ R ⁴ , -SR ³ , -SO, R ³ , -CO ₂ R ³ and -CONR ³ R ⁴ ;

(d) C, g-cycloalkyl, optionally substituted with 1 to 2 substituents independently selected from fluoro, chloro, bromo, -OR- ³ . -OCOR ³ , -OCONR ³ R ⁴ , oxo, -NR ³ R ⁴ , -NR ³ COR ⁴ , -NR ³ CONR ³ R,

-NR ³ SO ₂ R ⁴ , -SR ³ , -SO, R ³ , -CO ₂ R ³ and -CONR ³ R ⁴ ;

(e) cycloalkylalkyl of 3 to 6 carbon atoms in the cycloalkyl ring and 1 to 6 carbon atoms in the alkyl moiety, optionally substituted by 1 to 2 substituents independently selected from fluoro, chloro. Bromine, -OR ³ , -OCOR ³ , -OCONR ³ R ⁴ , oxo, -NR ³ R ⁴ , -NR ³ COR ⁴ , -NR ³ CONR ³ R ⁴ , -NR ³ SO ₂ R ⁴ , -SR ³ , -SO ₃ R ³ , -CO ₂ R ³ and CONR ³ R ⁴ ;

(f) heteroaryl, wherein the heteroatom or heteroatoms are selected from 1 to 4 oxygen, nitrogen or sulfur atoms, optionally substituted by 1 to 2 substituents independently selected from fluoro, chloro, bromo,

-OR ³ , -OCOR ³ , -OCONR ³ R ⁴ , oxo, -NR ³ R ⁴ , -NR ³ COR ⁴ , -NR ³ CONR ³ R ⁴ , -NR ³ SO ₅ R ⁴ , -SR ³ , -SO , R ³ , -CO ₅ R ³ and -CONR R; preferred heteroaryl radicals are 5- or 6-membered aromatic heterocyclic rings;

(g) heteroaralkyl wherein the heteroatom or heteroatoms are selected from 1 to 4 oxygen, nitrogen or sulfur atoms and the alkyl moiety has from 1 to 6 carbon atoms optionally substituted with 1 to 2 substituents independently selected from fluoro, chloro .

Bromine, -OR ³ . -OCOR ³ , -OCONR ³ R ⁴ , oxo, -NR ³ R ⁴ , -NR ³ COR ⁴ ,

-NR ³ CONR ³ R ⁴ , -NR ³ SO ₅ R ⁴ , -SR ³ , -SO ^ R ³ , -CO ₅ R ³ and

^ 4 -CONR R; preferred heteroaralkyl groups are those wherein the heteroaryl group is a 5- or 6-membered aromatic heterocyclic ring and the alkyl moiety has 1 to 2 carbon atoms;

(h) heterocyclyl, wherein the heteroatom or heteroatoms are selected from 1 to 4 oxygen, nitrogen or sulfur atoms, optionally substituted by 1 to 2 substituents independently selected from fluoro, chloro, bromo,

-OR ³ , -OCOR ³ , -OCONR ³ R ⁴ , oxo, -NR ³ R ⁴ , -NR ³ COR ⁴ , -NR ³ CONR ³ R ⁴ , -NR ³ SO ₄ R ⁴ -SR ³ , -SO _x R ³ , -CO ₀ R ³ and

X Λ ^ 2 cL

-CONR ^ R; preferred heterocyclyl groups are 5- or 6-membered, saturated or unsaturated rings;

(i) heterocyclylalkyl wherein the heteroatom or heteroatoms are selected from 1 to 4 oxygen, nitrogen or sulfur atoms and the alkyl moiety has 1 to 6 carbon atoms, optionally substituted with 1 to 2 substituents independently selected from fluoro, chloro , Bromine, -OR ³ . -OCOR ³ , -OCONR ³ R ⁴ , oxo, -NR ³ R ⁴ , -NR? COR ⁴ ,

-NR ³ CONR ³ R, -NR ³ SO ₄ R ⁴ , -SR ³ , -SO, R ³ , -CO ₀ R ³ and -CONR 1 R; preferred heterocyclylalkyl radicals are those wherein the heterocyclyl moiety is a 5- or 6-membered saturated or unsaturated ring.

ω Fluorine, chlorine or bromine; -OCONR ³ R; (K) -OR ³ ; -OSO2R ^3; (D -OCO ₀ R ³ oxo; (m) -OCOR ?; -NR ³ R ⁴ ; (Η) R ³ CONR ⁴ : (Ο) -NR ³ CO 2 R; , (P) -NR ³ CONR ³ R ⁴ ; (Q) -NR ³ SO ₂ R ⁴ ; (Γ) -SR ³ ; Cs) 0 (T) IQ C * Ό · (U) 0 0 (Ν) -S ^ R ⁹ ; ' -SO ₃ R ³ ; (W) -CO ₂ R ³ ; (Χ) (Y). (Ζ)

(aa) -CONR ³ R ⁴ ;

(bb) -CN; or (cc) phenyl, optionally substituted with 1-3 fluoro, chloro, bromo, C 1-6 alkyl, -OR ³ , -NR ³ R ⁴ , -SO ₃ R ³ , -CO ₂ R ³ or -CONR ³ R ⁴ .

The above substituents R ^3, R and R 1 • loading sitting in connection with the substituent R

given meanings

 The above-defined ring

-H.

is a non-aromatic, heterocyclic group. However, this ring may be fused to another ring which is a saturated or unsaturated carbocyclic ring, preferably a C._y-carbocyclic ring, a phenyl ring, a 4- to 7-membered heterocyclic (saturated or unsaturated) ring with 1 to 3 heteroatoms selected from O , N, S ( O ) _m , NR ^ and Λ7 Λ 8 NR R, or a 5- to 6-membered,

heteroaromatic ring containing 1 to 3 heteroatoms selected from O, S (O) _n , N, NR ¹⁵ and NR ¹⁷ R ¹⁸ , wherein m, R, R 'and R have the meanings given above

 30

λ c. Ί year

The substituent R of the non-aromatic radical R may be either (a) an optionally substituted C _1-6 alkyl, C ₂ _ ₁₀ alkenyl, C _2-1 alkynyl, C _3-6 cycloalkyl, C, _g -Cycloalkyl-C ₁ _g-alkyl 7 phenyl, phenyl-C _{1-> 6-} alkyl, phenyl-C ₂ _g-alkenyl, phenyl-C ₂ _g-alkynyl,

Heteroaryl, Heteroaralkyl-, wherein the alkyl moiety has 1 to 6 carbon atoms, heterocyclyl or HeteroeyeIylalkyl group in which the alkyl moiety has 1 to 6 carbon atoms, or (b) a divalent phenylene group or C ^^ - alkylene group, which at the ring

-N

is bound to form a ring-bridged, polycyclic group, e.g. a quinuclide group. The heteroaryl substituent (or the heteroaryl portion of the heteroaralkyl substituent) may be a mono-, M- or poly-ethylene, aromatic, heterocyclic group containing 1 to 4 O, N or S atoms; preferred are 5- or 6-membered heterocyclic rings such as thienyl, furyl, thiadiazolyl, oxadiazolyl, triazolyl, isothiazolyl, thiazolyl, imidazolyl, isoxazolyl, tetrazolyl, oxazolyl, pyridyl, pyrazinyl, pyrimidinyl, pyridazinyl, pyrrolyl and pyrazolyl. The heterocyclyl substituent (or the heterocyclyl portion of the heterocyclylalkyl substituent) may be a mono-, bi- or polycyclic, saturated or unsaturated non-aromatic heterocyclic group containing 1 to 4 O, N or S atoms; preferred are 5- or 6-membered heterocyclic rings such as morpholinyl, piperazinyl, piperidyl, pyrazolinyl, pyrazolidinyl, imidazolinyl, imidazolidinyl, pyrrolinyl and pyrrolidinyl.

If the substituent R ¹ is an alkyl, alkenyl, alkynyl, cycloalkyl, cycloalkylalkyl, phenyl, phenylalkyl, phenylalkenyl, phenylalkynyl, heteroaryl, heteroaralkyl, heterocyclyl or heterocyclyl

alkyl groups, these groups may be optionally substituted by 1 to 3 substituents selected from the following groups:

(a) C 1-6 alkyl optionally substituted by preferably 1 to 3 amino, fluoro, chloro, carboxyl, hydroxy or carbamoyl groups;

(b) fluorine, chlorine or bromine; (c) ^-OR3;

(d) -OCO ₂ R ³ ;

(e) -OCOR ³ ;

(f) -OCONR ³ R ⁴ ;

(g) -OSO ₂ R ³ ; (h) oxo;

(i) -NR ³ R ⁴ ; (j) R ³ CONR ⁴ : (k) -NR ³ CO 2 R; (1) -NR ³ CONR ³ R ⁴ ; (m) -NR ³ SO ₂ R ⁴ ; (n) -SR ³ ; (o) -SOR ⁹ ; Cp) -SO ₂ R ⁹ ; (q) -SO ₃ R ³ ; (r) -CO ₂ R ³ ;

(s) -CONR ³ R ⁴ ; (t) -CN; or

(u) phenyl, optionally substituted by up to 3 substituents independently selected from fluoro, chloro, bromo, C 1-10 alkyl, -OR ³ , -NR ³ R ⁴ , -SO, R ³ , -CO ₉ R ³ or CONR ³ R ^4, wherein in the Obi gen substituents R ^ID, the groups Rr and R are independently selected from hydrogen; Alkyl, alkenyl and alkynyl of 1 to 10 carbon atoms; Cycloalkyl, cycloalkylalkyl and AlkyleyeIoalkyl with 3

to 6 carbon atoms in the cycloalkyl ring and 1 to 6 carbon atoms in the alkyl moieties; phenyl; Aralkyl, aralkenyl and aralkinyl, wherein the aryl moiety is phenyl and the aliphatic moiety has from 1 to 6 carbon atoms; and heteroaryl, heteroaralkyl, heterocyclyl and heterocyclylalkyl, wherein the heteroaryl and heterocyclyl group or part of this group is as previously defined for R and the alkyl moieties attached to said heterocyclic moieties have from 1 to 6 carbon atoms; or

3 4

Br and R, together with the nitrogen atom to which at least one of these radicals is bonded, can form a 5- or 6-membered nitrogen-containing heterocyclic ring (as defined above for B ); and R ^{9 has} the meanings given above for Br , but can not stand for hydrogen. The most preferred substituent R is a C ₁ _g alkyl group, especially a methyl group.

1 6 If R is a divalent phenylene or C 1-6 alkylene

means group, then this group is to another atom of the ring of formula 25

^: N

bonded to form a bridged polycyclic ring, e.g. a quaternized quinuclidine ring of the formula

According to a particularly preferred embodiment, compounds of the general formula I are prepared according to the invention, wherein the radical of the formula

10 has the following meanings:

CH, .V

 Ν

N-CH.

CH.

S- »0

.CH.

CH-

or

wherein Y is a hydrogen, chlorine, bromine, iodine or fluorine atom or a C _1-6 -AlRyI-, hydroxy, -S-C ₁ _ ₆ ~ alkyl, carboxyl, carbamoyl or phenyl group. In this subclass, preferred compounds are those

wherein A is -CHCH ₂ -, - / \, -CH ₂ CH- or - (CH ₂ ) _n ,

CH ₃ > ~ ^ CH ₃ in which η is 2, 3 or 4, and especially those

wherein A is -CH ₂ CH ₂ -, -CH ₂ CH ₂ CH ₂ -, -CHCH ₂ -, - / \ or

CH ₃ ^

-CH ₉ CH-, and most preferably those wherein A is CH ₃

for -CH ₀ CH ₀ -, and in which either

1 8 (a) R and R together for

CH ₃

HIGH ₂

C =

CH ₃ stand or

CH ₃

(b) R is a hydrogen atom and R is for

CH, CH _x OH OH hydrogen, CH ₃ CH ₂ -, CH-, ^ C- or CH ₃ CH-

stands.

CH ₃ CH ₃

Especially preferred are compounds wherein R is for

OH

1 '

is a hydrogen atom and R is CH ₃ CH-, in particular, compounds of the absolute configuration 5R, 6S, 8R are preferred.

In a still more preferred embodiment

According to the invention compounds of general formula I prepared, wherein the radical of the general formula

"Ν

has the following meanings:

CH.

CH,

N-CH,

CH

3 \

-Ν

CH,

CH.

^c xT ~ \

CH.

-Ν '

NH or

In this preferred subclass are forthcoming

compounds in which A is -CHCH ₂ -, "" / - CH ₂ CH-

In this preferred subclass, preferred

CH, - "CH

- (CH ₂ ) - is, in which

such as those in which A is for -

-CHCHp-, - / N or -CHpOH-, and most preferably

^{stands for} 2, 3 or 4 before

CH,

CH

those in which A is -CHpCHp- and in which either

1 ß 30 (a) R and R together for

C =

stand or

(b) R is a hydrogen atom and R is for

CH ₃ CH ₃ OH OH

Hydrogen, CH ₃ CH ₂ -, CH-, ^ C or CH ₃ CH-

  CH, CH,

j j

stands.

• 8th

Particularly preferred are compounds wherein R is a hydrogen atom and R is for

Oh T

CH ^ CH- stands, in particular compounds of the absolute configuration 5R, 6S, 8R.

In a further, more preferred embodiment, compounds of the general formula I are prepared according to the invention, wherein the rest of the formula

For

CH,

In which Y is a hydrogen, chlorine, bromine, iodine or fluorine atom or a C, g-alkyl, hydroxy, -SC ₁ _g-alkyl, carboxyl, carbamoyl or phenyl group. In this preferred subclass, the preferred compounds are those wherein A is "" (CHp) "" wherein η is 2, 3 or 4. Most preferred are those compounds wherein A is and wherein either

(a) R and R together for

^l 2 ^

CH,

stand or

(b) R is a hydrogen atom and R is CH ₃ CH ₃ OH OH hydrogen, CH ₃ CH ₂ -, CH-, ^ C- or CH ₃ CH-

CH,

CH,

stands.

Especially preferred are compounds wherein R is an OH

1 ♦

Is hydrogen and R is CH ^ CH-, in particular compounds of the absolute configuration 5R, 6S, 8R.

In a most preferred embodiment according to the invention compounds of the general formula I are obtained, wherein the radical of the formula

25

For

30

^ O

stands.

35

In this preferred subclass preferred compounds are those wherein A is - (CHp) -, where η is 2, 3 or 4, and most preferably those wherein A is -CH ₉ CH ₀ - and wherein either

18

(a) R and R together for

HIGH ₂

C = CH,

stand or

Q A

(b) R is a hydrogen atom and R is for

CH ₃ CH ₃ OH OH

Hydrogen, CH ₃ CH ₂ - CH-, Z- or CH ₃ CH-15 ^ CH ₃ ^ CH ₃

stands.

Especially preferred are compounds wherein R is a

OH

1 t

Is hydrogen and R is CH, CH-, especially compounds of absolute configuration 5R,. 6S, 8R. '

In a most preferred embodiment, compounds of the general formula are obtained according to the invention

OH ^H " _R 16.

(R)

-S-CH ₂ CH ₂ -N

^ '/ ^ COOR

in which - ^

For

N-CH ₃ or

(both α- and β-diastereoisomers)

stands

and wherein R represents a hydrogen atom, an anionic charge or a common, easily removable carboxyl-protecting group, provided that, if R represents a hydrogen atom or a protecting group, there is also a counterion, and the pharmaceutically acceptable addition salts thereof.

Certain compounds covered by general formula I may exist both as optical isomers and as epimer mixtures. According to the invention, all of these optical isomers and the epimer mixtures are included. For example, if the substituent in the 6-position is a hydroxyethyl group, then this substituent may be present in both the R and S configurations. Thus, according to the invention, both the obtained isomers and their epimer mixtures are included.

In the process according to the invention, an intermediate compound of the general formula IV is used

"COOR '

IV

a, which is disclosed for example in European Patent Application 38 869 and which can be prepared by the general method described therein. X denotes a customary leaving group (it is designated "X" in European Patent Application 38 869), such as chlorine, bromine, iodine, benzenesulfonyloxy, p-toluenesulfonyloxy, p-nitrobenzenesulfonyloxy, methanesulfonyloxy, trifluoromethanesulfonyloxy, diphenoxyphosphinyloxy or di (trichloroethoxy) -phosphinyloxy. The preferred leaving group is diphenoxyphospinyloxy. The intermediates of general formula IV are generally formed in situ by reaction of an intermediate compound of general formula III

2 '

III

1 8 2 ^f

wherein R, R and R have the meanings given above, with a suitable acylating agent R -L. The preferred intermediate IV, in which L is a diphenoxypho sphinyloxy group, can be prepared by

len, by reacting a ketoester of formula III in an inert organic solvent such as methylene chloride, acetonitrile or dimethylformamide, with about one molar equivalent of diphenylchlorophosphate in the presence of a base such as diisopropylethylamine, triethylamine, 4-dimethylaminopyridine or the like., At a temperature of about -20 ° to + 40 ⁰ C, most preferably at about 0 ° C, reacted. The intermediate compound IV can be isolated if desired; However, it is usually used without further isolation or purification as a starting compound for the process according to the invention.

In the process according to the invention, a carbene penem compound IV is used with a quaternary amine thiol compound of the general formula VII

HS-A-R ¹⁴ X

VII

wherein A is a cyclopentylene, cyclohexylene or Cpr-alkylene group, which is optionally substituted by one or more C ^ _ _{Z +} alkyl groups, most preferably 5 for a cyclopentylene group, a cyclohexylene group or

R ¹⁰ R ¹²

I i

_C C-

»»

in which R ¹⁰ , R ¹¹ , R and R ¹ 'each independently of one another represent a hydrogen atom or a C 1-10 -alkyl group, X represents a counterion which originates from a strong acid, eg Cl -, Br ",

CH ₃ SO ₃ ", CF ₃ SO ₃ " or CH ₃ - ^ V) -SO ₃ ", and R ¹ ^ a

as defined above, quaternized, nitrogen-containing, aromatic or non-aromatic heterocycle. The reaction is carried out in an inert solvent, such as acetonitrile, acetonitrile-dimethylformamide, tetrahydrofuran, tetrahydrofuran-HpO, acetonitrile-H ₂ O or acetone, in the presence of a base. The type of base is not critical. However, the best results are obtained when using a non-nucleophilic tertiary amine as the base, such as diisopropylethylamine, 1,8-diazabicyclo [5.4.0] undec-7-ene, 1,5-diazabicyclo [4.3.0] - non-5-ene, or a Trl (C ₁ _ _> Zf) alkylamine, such as triethylamine, tributylamine or tripopylamine. The reaction of the intermediate compound rv with the thiol VII can be in a wide temperature range, at -15 ⁰ C up to room temperature, for example, carry. Preferably, one operates in a temperature range of about -15 ° to +15 ⁰ C, most preferably about ⁰ ° C.

The carbapenem compound obtained by reacting the quaternary amine of Formula VII with the intermediate IV has a counter anion [eg (CgH ₅ O) ₂ PO ₂ ⁶ , Cl ⁹ or the quaternary thiol associated anion] associated with the compound. This counteranion can be replaced at this stage by conventional methods with another counteranion, for example, one that is more pharmaceutically acceptable. Alternatively, one can remove the counter anion during the subsequent deblocking step. When the quaternized carbapenem compound and the counter anion form an insoluble product, the product can be allowed to crystallize out as it is formed and can be recovered by filtration.

After obtaining the desired carbapenem compound

2 ', one can, if desired, remove the carboxy-1 protective group R of the compound I ¹ by customary processes, such as solvolysis, chemical reduction or hydrogenation. When the protecting group is a p-nitrobenzyl, benzyl, benzhydryl or 2-naphthylmethyl group which is readily removed by catalytic hydrogenation, the intermediate compound I ^{1 is} treated in a suitable solvent such as dioxene-water-ethanol , Tetrahydrofuran-diethyl ether buffer ,. Tetrahydrofuran-aqueous .Dikaliumhydrogenphosphat-isopropanol or the like., At a hydrogen pressure of 1 to 4 at in the presence of a hydrogenation catalyst such as palladium-on-carbon, palladium hydroxide, platinum oxide or the like, at a temperature of 0 to 50 ⁰ C during about

0.24 to 4 hours. If the group R is o-nitrobenzyl, then one can use a photolysis for deblocking. Protecting groups, such as 2,2,2-trichloroethyl, can be removed by mild reduction with zinc. The allyl protecting group can be removed by using a catalyst such as a mixture of a palladium compound and triphenylphosphine in a suitable aprotic solvent such as tetrahydrofuran, methylene chloride or diethyl ether. Similarly, one can remove other common carboxy1 protecting groups by methods known to those skilled in the art.

Finally, one can, as already stated above, the

2 ¹

Compounds of formula I ^f , wherein R represents a physiologically hydrolyzable ester group, such as acetoxymethyl, phthalidyl, indanyl, pivaloyloxymethyl, methoxymethyl, etc., directly to the host without deblocking, since such esters are hydrolyzed in vivo under physiological conditions.

1 ft

If the substituent R and / or R or the quaternized nucleos attached to the substituent A contain

14

If phil R is a functional group that might interfere with the intended course of the reaction, then this group can be protected with a common blocking group and then deblocked to regenerate the desired functional group. Suitable blocking groups and methods for introducing and removing such groups are well known to those skilled in the art.

Like the other β-lactam antibiotics, the compounds of the general formula I can be converted by known methods into pharmaceutically acceptable salts, which for the purposes according to the invention are essentially equivalent to the compounds not converted into the salts. So you can, for example, a

2 Dissolve the compound of the general formula I in which R is an anionic charge in a suitable inert solvent. Then one adds one equivalent of a pharmaceutically acceptable acid. The desired acid addition salt can be prepared by conventional methods, e.g. Solvent precipitation, lyophilization, etc., win. If other basic or acidic functional groups are present in the compound of general formula I, the pharmaceutically acceptable base addition salts and acid addition salts can be obtained in a similar manner by known methods. j

^r Second

The compounds of the general formula I in which R is a hydrogen atom or an anionic charge, or pharmaceutically acceptable salts thereof, can also be converted into the corresponding compounds by conventional methods.

convert compounds in which R is a physiologically hydro-

lysable ester group means. Compounds of general

P my formula I, wherein R is a common carboxyl-protecting group, can be in the corresponding

P bonds, where R is a hydrogen atom, an anionic

Means charge or a physiologically hydrolyzable ester group, or convert into the pharmaceutically acceptable salts thereof. Some of the thiol intermediates of general formula VII may be prepared by, for example, adding a sulfide of the formulas

or

VIIIb

Wherein R, R, R and R 1 each independently represent a hydrogen atom or a C 1-4 alkyl group, with a heteroaromatic amine (as defined above) of the general formula

or with a non-aromatic, heterocyclic amine (as defined above) of the general formula

R ^1S

and reacted with a strong acid. The reaction can be carried out in the presence or absence of an inert organic solvent. This solvent is preferably a non-polar, organic solvent such as methylene chloride, benzene, xylene, toluene or the like. If the amine and sulfide reagents are liquids or a solid amine is soluble in a liquid sulfide reagent, then the reaction is preferably carried out without additional Solvent through.

The strong acid used for the reaction is not critical and may be, for example, a strong inorganic or organic acid such as hydrochloric acid, hydrobromic acid, methanesulfonic acid, p-toluenesulfonic acid, trifluoromethanesulfonic acid, etc.

The formation of the quaternary Aminthiol intermediate of formula VII can be carried out in a temperature range of about -20 ° to about 100 ⁰ C. Preferably, one works at temperatures of about 50 to 70 ⁰ C.

The sulfide reagent, the aromatic amine and the acid are preferably used such that the sulfide and acid are present in approximately equimolar amounts, with the amine being used in excess, e.g. 2 to 3 moles of amine per mole of sulfide or acid.

The quaternary Aminthiol intermediate has an associated counter anion derived from the acid used. It is of course possible

replace the anion by another Gegenanion by conventional methods, which is then used in the subsequent reaction with the carbapenem intermediate IV.

The carbapenem derivatives of general formula I, wherein

ρ R represents a hydrogen atom, an anionic charge or a physiologically hydrolyzable carboxyl-protecting group, or the pharmaceutically acceptable salts thereof are effective antibiotics which can be used against various Gram-positive and Gram-negative bacteria. They may also be used, for example, as animal feed supplements for increasing vegetable growth, as food preservatives, as bactericides for industrial applications, e.g. in water-based paints or in the washing water of paper mills for inhibiting the growth of dangerous bacteria, and as a disinfecting agent for destroying or inhibiting the growth of dangerous bacteria on medical and dental equipment. However, they are particularly useful for treating infectious diseases caused by Gram-negative or Gram-positive bacteria in humans and animals.

The pharmaceutically active compounds provided by the invention may be used alone or as pharmaceutically-formulated agents. Such agents contain, in addition to the active carbapenem compound, a pharmaceutically acceptable carrier or diluent. The compounds can be administered in a variety of ways. These include in particular the oral, topical or parenteral (intravenous

or intramuscular injection) administration. The pharmaceutical agents may be in solid form, e.g. as capsules, tablets, powders, etc., or in liquid form as solutions, suspensions or emulsions. Injection agents (preferred mode of administration) may be prepared in unit dosage form in ampoules or may be in multi-dose containers and may contain formulatory agents such as suspending agents, stabilizing agents or dispersing agents. The agents may be in ready to use form. They may also be in powder form, such that at the time of administration they may be supplemented with a suitable carrier, e.g. sterile water, reconstituted.

The dose to be administered will depend primarily on the compound used, the specific formulation, the mode of administration, the nature and condition of the host. Of course, the dosage also depends on the specific situs and the organism to be treated. The choice of the most preferred dosage and the route of administration is left to the treating physician. In general, however, the compounds are administered parenterally or orally to a mammal (human and animal) in an amount of about 5 to 200 mg / kg / day. Administration is generally in several doses, e.g. three or four times a day.

The following examples serve to illustrate the invention.

example 1

Preparation of 3- (2- (1-pyridinium) ethylthio) -6a (1 (R) -hydroxyethyl) -7-OXO-1-azabicyclo [3.2.0] hept-2-ene

2-carboxvlat

A. 1- (2-Mercaptoethyl) pyridinium methanesulfonate

xSv VN \ ^ ^ N (+)> MsO

55 C, 16 h

A suspension of pyridinium methanesulfonate in pyridine is prepared by adding dropwise methanesulfonic acid (1.95 ml, 0.03 mol) to pyridine (8.0 ml, 0.099 mol) with cooling. To this suspension is added ethylene sulfide (1.96 ml, 0.033 mol). The resulting mixture is stirred for 16 h at 55 ° C, concentrated under reduced pressure to a thick syrup, which is mixed with a few ml of water and the solution is poured onto a column (40 χ 16 cm) of / u Bondapak C-18 which is eluted with water. Lyophilization of the appropriate fractions results in a colorless syrup; Yield 6.5 g (91%).

IR (movie) ν? : 2300-2600 (br, SH), 1635 (pyridinium), 1490, 1200 (sulfonate), 1068, 1060, 1045, 791, 780 cm ^{-1 1} H-NMR (DMSO-d ₆ ) 6 : 2.32 ( 3H, s, CH ₃ SO ₃ "), 2.61, 2.70, 2.73, 2.82 (1H, B portion of the A ₂ B system, SH), 3.07 (2H, m [ with D ₂ O: 3.08 (2H, t, J = 6.5 Hz)], CH ₂ S), 4.76 (2H, t, J = 6.5 Hz, CH ₂ N ⁺ )., 8 , 19 (2H, m, Hm of pyridinium), 8.6 (1H, m, Ho of pyridinium), 9, L8 (2H, dd, J = 6.8 Hz, J = 1.4 Hz, Ho of pyridinium )

UV (H ₂ O)

5 Method A

: 206 (6 5230), 258 (£ 3760) ni / U

For example, 1- (2-mercaptoethyl) -T3vridiniiimchlorid

-MsO

Permutit S-1

cl "

An aqueous solution of crude 1- (2-mercaptoethyl) -pyridinium methanesulfonate (9.4 g, 0.04 mol) is added to a column (2.5 x 41 cm) of Permutit S-1 Cl " Column is eluted at a rate of 0.5 ml / min, the appropriate fractions combined and lyophilized to give a yellow syrup, yield 7.0 g (100%), which is thus used for the next step.

¹ H-NMR (D ₂ O) S : 3.22 (2H, m, CH ₂ S), 4.88 (m, CH ₂ N ⁺ ), 8.18 (2H, m, Hm of pyridinium), 8 , 7 (IH, m, Hp of pyridinium), 9.0 ppm (2H, m, Ho of pyridinium).

25 Procedure B

Cl

To pre-cooled (ice bath) pyridine (5.6 mL, 70 mmol) is added pyridine hydrochloride (4.05 g, 35 mmol) and ethylene sulfide (2.1 mL, 35 mmol). The mixture is warmed to 65 ° C and stirred for 75 min, resulting in a two-phase system

receives. The lighter phase is removed. The remaining oil is washed with ether (5x10 ml) and a high vacuum is applied to give the title compound (90-100%) which is then used for the next step.

C. p-Nitrobenzyl-3- [2- (1-pyridinium) ethylthio] -6a [i-

(R) -hydroxyethyl] -7-oxo-1-azabicyclo [3.2.0] hept-2- ene-2-carboxylate chloride

OH

1) NEt (IPr) ₀ 0 ^

Z) ClP (OPh) ₂

COOPNB 4) NEt (IPr) ₂

COOPNB

A solution of p-nitrobenzyl 6α- [i- (R) -hydroxyethyl] -3,7-dioxo-1-azabicyclo [3.2.0] heptane-2-carboxylate (6.09 g, 17.5 mmol) in acetonitrile (20 ml) is cooled under nitrogen atmosphere to +5 ⁰ C, treated sequentially with diisopropylethylamine (3.65 ml, 21.0 mmol) and diphenyl chlorophosphate (4.34 ml, 21.0 mmol), the mixture obtained is stirred for 30 min at 5 ° C _y-cooled to -5 ° C and treated sequentially with a solution of crude 1- (2-mercaptoethyl) pyridinium chloride (4.3 g, 24 mmol) in N, N-dimethylformamide (1.0 ml) and dropwise with diisopropylethylamine (3.65 mL, 21.0 mmol). The reaction mixture is stirred for 1 h at ^{0 0} C, cooled to -30 ⁰ C and stirred for a further 15 min. The solid is filtered off and washed with cold (-30 ⁰ C) acetonitrile to give 5.77 g (65%) yield.

IR (nujol) 0 ⁾ _max : 3300 (OH), 1775 (C = O of β-lactam), 1690 (C = O of PNB ester), 1630 (pyridinium), 1605 (phenyl of PNB ester), 1515 (NO ₂ ),

1335 cm ' ¹

¹ H-NMR (DMSO-d ₆ ) 6 : 1.17 (3H, d, .J = 6.1 Hz, CH ₃ CHOH), 3.2-3.75 (5H, H-4, H-6 , CH ₂ S), 3.75-4.5 (2H, H-5, CH ₃ CHOH), 4.92 (2H, br.t, J = 6.5 Hz, CH ₂ N ⁺ ), 5, 18 (1H, d, J = 4.9 Hz, OH), 5.37 (center of the ABq, J _ab = i4.2 Hz, CH ₂ of PNB), 7.69 (2H, d, J = 8, 7 Hz, Ho of PNB), 8.24 (d, * J = 8.7 Hz, Hm of PNB), 8.0-8.4 (4H, Hm of PNB, Hm of pyridinium), 8.66 ( 1H, m, Hp of pyridinium), 9.17 (2H, br.d, J = 5.5 Hz, Ho of pyridinium).

The filtrate and the solvent used for washing are combined and diluted with 150 ml of ether. The supernatant liquid is decanted and the gum product dissolved in 40 ml of water containing acetonitrile sufficient to give a solution which is applied to a column (3 x 10 cm) of / U Bondapak C-18. The column is eluted with 10% acetonitrile-90% water (150 ml) and 50% acetonitrile-50% water (100 ml) mixtures. The appropriate fractions are combined and lyophilized after removing the acetonitrile in vacuo. A yellow powder is obtained. NMR shows the presence of the title compound which is treated with some p-nitrobenzyl-3- [2- (1-pyridinium) -ethylthio] -6cc- [i - (R) -hydroxyethyl] -7-oxo-1-azabicyclo [ 3.2.0] hept-2-ene-2-carboxylate diphenyl phosphate (2: 1). Dissolve the powder in as little water as possible and add water over a column (1.5 x 21 cm) of permutite S-1. "Lyophilization of the appropriate fractions gives 1.8 g (20%) of the title compound ,

D. p-Nitrobenzyl-3- [2- (1-pyridinium) -ethylthio] -6oc- [1- (R) -hydroxyethyl] -7-oxo-1-azabicyclo [3.2.0] hept-2-ene 2-carboxylate dipheny! phosphate

OH 1) NEt (IPr) ₉

I 2

^ "'- ^ 2) ClP (OPh)

N,

COOPNB

4) NEt (IPr) ₂

A solution of ρ-Ni-trobenzyl-6oc- [i- (R) -hydroxyethyl] -3,7-dioxo-1-azabicyclo [3.2.0] heptane-2-carboxylate (0.174 g,

* 15 0.50 mmol) in acetonitrile (2 ml) is cooled under nitrogen atmosphere to 0 ° C, treated successively with diisopropylethylamine (0.105 ml, 0.60 mmol) and diphenylchlorophosphate (0.124 ml, 0.60 mmol), the stirred solution obtained at ⁰ ° C. for 30 minutes and treated successively with a solution of 1- (2-mercaptoethyl) -pyridinium methanesulfonate (0.170 g, 0.72 mmol) in 0.6 ml of acetonitrile and diisopropylethylamine (0.105 ml, 0.60 mmol). The reaction mixture is stirred for 15 min at ⁰ ° C. Dilute with cold (0 ° C) water (7 mL) and place the mixture on a column (1.5 x 6.4 mm) of / U Bondapak C-18. The column is eluted with a mixture of acetonitrile (25% -50%) in water (75% -50%). The appropriate fractions are combined and lyophilized after removing the acetonitrile in vacuo. You get a yellow

30 powders; 0.33 g (92%).

IR (KBr)% 7 _max : 3600-3000 (OH), 1765 (C = O of β-lactam), 1690 (C = O of PNB ester), 1625 (pyridinium), 1585 (phenyl), 1510 (NO ₂ ), 1330 (NO ₂ ), 885 cm ^-1 (NO ₂ )

¹ H-NMR (DMSO-d ₆ ) 6 : 1.16 (3H, d, J = 6.2 Hz, CH ₃ CHOH), 4.87 (2H, br.t, J = 6.6 Hz, CH ₂ S), 5.37 (center of ABq, J -, = 14.3 Hz, CHo of PNB), 6.7-7.5 (phenyl), 7.68 (d, J = 8.8 Hz, Ho of PNB), 8.23 (d, J = 8.8 Hz, Hm of PNB), 8.0-8.3 (m, Hm of pyridinium), 8.4-8.8 (1H, Hp of Pyridinium), 9.09 (2H, dd, J = 6.7 Hz, J = 1.3 Hz, Ho of pyridinium).

E. 3- [2- (1-Pyridinium) ethylthio] -6a [i (R) -hydroxyethyl J-7-OXO-1-azabicyclo [3.2.0] hept-2-ene-2- carboxylate Method A

25

OH

20 ο

TQ

% Pd / C, H ₂ THF, ether

COOPNB

(PhO) ₉ PO '

COO

To a solution of p-nitrobenzyl-3- [2- (1-pyridinium) ethylthio] -6 <x- [i - (R) -hydroxyethyl] -7-oxo-1-azabicyclo [3.2.0] hept. Add 2-ene-2-carboxylate-diphenylphosphate (0.16 g, 0.22 mmol) in humid tetrahydrofuran (10 mL) to ether (10 mL). Potassium phosphate monobasic sodium hydroxide buffer pH 7.4 (16 mL). 0.05 M) and 10% palladium on carbon (0.16 g). The mixture obtained is hydrogenated for 1 h at 25 ° C and 2.76 bar (40 psi). Man.trennt the

two phases and the organic phase is washed with water (2x3 ml). The aqueous solutions are combined, washed with ether (2x10 ml) and added to a column (1.5 x 6.2 cm) of / U Bondapak C-18 after removing traces of the organic solvents in vacuo. The column is eluted with water and, after lyophilization of the appropriate fractions, a yellow powder is obtained, yield 0.062 g (84 #).

IR (KBr) \? _max : 3700-3000 (OH), 1755 (C = O of β-lactam),

1630 (pyridinium), 1590 cm ^-1 (carboxylate) ¹ H-NMR (D ₂ 0) <i: 1.22 (3H, d, J = 6.4 Hz, CH, CHOH), 2.92 (d, J = 9.1 Ηζ, Ή-4), 2.97 (d, J = 9.1 Hz, H-4), 3.20 (dd, J = 2.5 Hz, J = 6.1 Hz, H-6), 3.44 (t, J = 6.0 Hz, CH ₂ S), 3.93 (dd, J = 9.1 Hz, J = 2.5 Hz, H-5), 4, 82 (t, J = 6.0 Hz, CH ₂ N ⁺ ), 8.04 (m, Hm of pyridinium), 8.5 (m, Hp of pyridinium), 8.82 (dd, J = 3.2 Hz, J = 1.1 Hz, Ho of pyridinium) UV (H ₂ O) λ _max : 259 (£ 5800), 296 ( t 7O 3 O) _{m /} u

1.2

= 13.5 h (measured at a concentration of 10 ^"4 M in phosphate buffer pH 7.4 at 36.8 ⁰ C).

Method B

10% Pd / C, H _2, THF, ether, buffer 7.2

COO

To a solution of p-nitrobenzyl-3-C2- (1-pyridinium) -ethylthio] -6a - [- 1- (R) -hydroxyethyl J-7-0X0-1-azabicyclo [3.2.O] hept-2 -en-2-carboxylate chloride (5.77 g, 11.4 mmol) in potassium phosphate monobasic sodium hydroxide buffer (170 mL, 0.2 M, pH 7.22) are added tetrahydrofuran (30 mL), ether (30 ml) and 10% palladium on carbon (5.7 g), the resulting mixture hydrogenated at 22 ⁰ C and 40 psi for 1 h and filtered through a pad of celite. Wash the pad with water (2 x 15 ml), combine the filtrate and the washings and dilute with ether (100 ml). The aqueous phase is separated off, washed with ether (3 x 100 ml) and poured onto a column (4.5 x 20 cm) of / u-Bondapak C-18 after removing the organic solvents in vacuo. The column is eluted with water and then with a mixture of Λ% acetonitrile in water. After lyophilization of the appropriate fractions, 2.48 g (6596 g) of the title compound are obtained as a yellow powder. The analytical data are identical to those for the compound prepared according to Method A.

At a_v ie 1 2

Preparation of 3- [2- (1- (3,5-dimethylpyridinium) ethylthiol] -6a [1- (R) -hydroxyethyl] -7-oxo-1-azabicyclo [3.2.01- hept-2-ene -2-carboxylate '

COO

A. 1- (2-Mereaptoethyl) -3,5-dimethylpyridinium methanesulfonate

MsO "+ MsOH

To a suspension of 3,5-lutidinium methanesulfonate in 3,5-lutidine prepared by adding methanesulfonic acid (0.65 ml, 0.010 mol) to cold 3,5-lutidine (2.51 ml, 0.022 mol), Add ethylene sulfide (0.655 mL, 0.011 mol), stir the resulting mixture at 55 ° C under a nitrogen atmosphere for 24 h, cool to 23 ⁰ C and dilute with water (5 mL) and ether (5 mL). The organic layer is separated off and the aqueous solution is washed with ether (6 × 4 ml). The traces of ether are removed in vacuo and the solution placed on a column (2.5 x 6.0 cm) of / u-Bondapak C-18. The column is eluted with water and lyophilizes the appropriate fractions to give a colorless syrup, yield 2.4 g (91%).

IR (film) v) 2520 (SH), 1628 (pyridinium), 1600, 1495, 1325, 1305, 1283, 1200 (sulfonate), 1040, 938,

765, 680 cm ^{-1 1} H-NMR (DMSO-d ₆ ) S : 2.31 (3H, s, CH ₃ SO ₃ "), 2.47 (6H, s,

CH ₃ of pyridinium), 2.57, 2.66, 2.69, 2.78 (1H, B part of the A ₂ B system, SH), 3.06 (2H, m [with added D ₂ O (2H, t, J = 6.5 Hz)], CH ₂ S), 4.65 (2H, t, J = 6.5 Hz, CH ₂ N ⁺ ), 8.34 (1H, s, Hp of Pyridinium), 8.79 (2H, s, Ho of pyridinium) UV (H ₂ O) λ _max : 271 (£ 4860) nyu

Analysis: for C ₁ QH ₁₇ NO ₅ S ₂ * O, ₂

Calculated: C 44.09% H 6.66% N 5.14% S 23.54% Found: 44.26 6.49 5.17 24.18.

B. p-Nitrobenzyl-3- [2- (1- (3,5-dimethylpyridinium)) ethylthio] -6a- [1- (R) -hydroxyethyl] -7-oxo-1-azabicyclo- Γ3.2.0] hept-2-ene-2-carboxvlat-diphenvlphosphat

OH 1) NEt (IPr) ,,

O *

2) ClP (OPh) ₂

3)

COOPNB

4) NEt (iPr) ₂

(1- (R) -hydroxyethyl) To a cold ⁽⁰ ° C) solution of p-nitrobenzyl 6a- -3,7-dioxo-1-azabicyclo [3.2.0] heptane-2-carboxylate (0.523 g, 1 , 50 mmol) in acetonitrile (6.0 ml) are added under nitrogen atmosphere diisopropylethylamine (0.314 ml, 1.8 mmol) and then diphenylchlorophosphate (0.373 ml, 1.8 mmol). The reaction mixture is stirred for 30 minutes and treated with a solution of 1- (2-mercaptoethyl) -3,5-dimethylpyridinium methanesulfonate (0.493 g, 1.87 mmol) in acetonitrile (1.9 mL) followed by diisopropylethylamine (0.314 ml, 1.8 mmol). The reaction mixture is stirred for 1 h at 0 ° C, diluted with cold (0 ° C) water (26 ml) and poured onto a column (7.0 χ 3.5 cm) of / u-Bondapak C-18. The column is eluted with a 25-50% acetonitrile 75-50% water mixture and, after lyophilization of the appropriate fractions, 1.01 g (90%) of the titer compound is obtained as a yellow powder.

10

IR (KBr) 0 ! 3700-3100 (OH), 1778 (C = O of β-lactam), 1700 (C = O of the PNB ester), 1635 (pyridinium), 1595 (phenyl), 1521 (NO ₂ ), 1335 (NO ₂ ) , 895 cm " ¹

¹ H-NMR (DMSO-d ₆ ) 6 : 1.16 (3H, d, J = 6.1 Hz, CH ₃ CHOH), 2.43 (s, CH, of pyridinium), 4.75 (2H, m, CH ₂ N ⁺ ), 5.38 (center of ABq, J _b = 1.4 Hz, CH ₂ of PNB), 6.6-7.5 (1 OH, m, phenyl ) , 7.70 ( 2H, d, J = 8.7 Hz, Ho of PNB), 8.0-8.5 (3H, m, Hp of pyridinium, Hm of PNB), 8.82 (2H, s, Ho of pyridinium),

UV (H ₂ O) A

270 (E-11570), 306 (£ 7343)

¹ Z ' ' max ¹ analysis: for C

Calculated: C 58.03% H 5.26% N '5.48% S 4.18% Found: 57.98 5.05 5.22 4.34.

C. 3- [2- (1- (3,5-Dimethylpyridinium)) ethylthio] -6a [i - '(R) -hydroxyethyl] -7-oxo-1-azabicyclo [3.2.0] hept-2 - en-2-carboxvlate

10% Pd / C, H ₂

THF, ether,

buffer

30

COO

35

To a solution of p-nitrobenzyl-3- [2- (i- (3,5-dimethylpyridinium)) ethylthio] -6a- [i- (R) -hydroxyethyl) -7-oxo-5 '1-azabicyclo [ 3.2.0] hept-2-ene-2-carboxylate diphenyl phosphate (0.600 g, 0.80 mmol) in humid tetrahydrofuran (36 mL) is added ether (36 mL), potassium phosphate monobasic sodium hydroxide buffer (0.05 M , pH 7.4, 44 ml) and 10% palladium on carbon (0.60 g), the resulting mixture hydrogenates for 1.25 h at 23 ⁰ C and 2.76 bar (40 psi), separates the organic layer and extracted with the buffer (2x5 ml). The layers of water are combined, filtered through a celite pad, washed with ether (40 ml) and traces of the organic solvents removed in vacuo.

Then place the solution on a column (2.5 x 10.0 cm) of / u-Bondapak C-IB. The column is eluted with water and the appropriate fractions are lyophilized to give 0.1.86 g (64%) of the title compound as a yellowish powder. '

IR (KBr) v) _m : 3700-3100 (OH), 1760 (C = O of β-lactam), max λ

1595 cm (carboxylate

¹ H-NMR (D ₂ O) S : 1.21 (3H, d, J = 6.3 Hz, CH ₃ CHOH), 2.45 (6H, s, CH ₃ on pyridinium), 2.81 (i.e. , J = 9.2 Hz, H-4), 2.96 (d, J = 9.2 Hz, H-4), 3.22 (dd, J = 2.6 Hz, J = 6.2 Hz, H-6), 3.40 (t, J = 6.2 Hz, CH ₂ S), 3.84

(dd, J = 9.2 Hz, J = 2.6 Hz, H-5), 4.15 (m, CH ₃ CHOH), 4.71 (t, J = 6.2 Hz, CH ₂ N ⁺ ), 8.21 (1H, s, Hp of pyridinium), 8.46 (2H, s, Ho of pyridinium) UV (H ₂ O) \ _max : 279 (ε 8345), 296 (S7714) nyu [α] β ³ = +40.7 (c O> 53, H _£ 0)

Tw ₂ = 16.9 (measured at a concentration of 10 ~ ⁴ in phosphate buffer pH 7.4 at 36.8 ⁰ C) h.

Example 5

Preparation of (5R, 6S) -3 - [[2- (3-hydroxymethylpyridinio) ethyl] thio] -6- [i- (R) -hydroxyethyl] -7-oxo-1-azabicyclo [5.2.2] lhept -2-ene-2-carboxylate

CHoOH ι

A. 3-Hydroxymethyl-1- (2-mercaptoethyl) -pyridinium trifluoromethanesulfonate

CH ₂ OH

HSCH ₂ CH ₂

CH ₂ OH

Trifluoromethanesulfonic acid (1.327 ml, 0.015 mol) is added dropwise to 3-pyridinemethanol (2.91 ml, 0.030 mol) and then ethylene sulfide (0.89 ml, 0.015 mol) is added. The resulting homogeneous mixture is heated (oil bath) under N ₂ for 20 h at 50 to 70 ⁰ C, then the reaction mixture in water (15 ml) and extracted with CH ₂ Cl ₂ (5 x 5 ml). The aqueous phase is concentrated in vacuo and then placed on a CQ reverse phase column. It is eluted with water and then concentrated the appropriate fractions, giving a pale yellow oil. This material is chromatographed again to give an almost colorless oil. After being in

Vacuum (P ₂ Oc), the product is obtained (4.50 g, 9h%) as a viscous oil.

IR (FiIm) O _1021x : 3450 (st, OH), 2560 (schw, SH) cm ^{-1 1} H-NMR (dg-acetone) 6 : 9, 10-8.05 (m, 4H, aromatic), 5 , 01 (t, J = 5.5 Hz, 2H, N-CH ₂ ), 4.93 (s, 2H, -CH ₂ OH), 4.43 br.s, 1H, -OH), 3.43 -3.18 (m, 2H, S-CH ₂ ), 2.34-2.10 (m, 1H, SH). 10

B. p-Nitrobenzyl- (5R, 6S) -3- [2- (3-hydroxymethylpyridinio) ethylthioj-o-ti -] - hydroxyethylj ^ -oxo-i-azabicyclo-Γ3.2.01hept-2-en-2 carboxylate diphenylphosT3hat

CO ₂ PNB

CH ₂ OH

CO ₂ PNB

To a solution of p-nitrobenzyl- (5R, 6S) -6- [1- (R) -hydroxyethyl] -3,7-dioxo-1-azabicyclo [3.2.0] heptane-2-carboxylate (0.174 g, 0 , 50 mmol) in 2 ml of dry acetonitrile are added at ⁰ ° C. under N ₂ diisopropylethylamine (0.096 ml, 0.55 mmol), then diphenylchlorophosphate (0.114 ml, 0.55 mmol) is added dropwise and the reaction mixture is stirred for 30 min O ⁰ C. A solution of 3-hydroxymethyl-1- (2-mercaptoethyl) pyridinium trifluoromethanesulfonate

(0.233 g, 0.70 mmol) in 0.50 ml of acetonitrile are then added, followed by diisopropylethylamine (0.122 ml, 0.70 mmol), the mixture is kept for 30 min at ^{0 0} C and then concentrated the reaction mixture in vacuo. The remaining, yellow, rubbery residue is taken up in water, adding so much acetonitrile that the rubbery product goes into solution. This solution is applied to a C 2 -C 4 reverse phase column which is eluted with 15% acetone / tril water. The appropriate fractions are lyophilized to give the product (0.305 g, 81%) as a beige solid.

IR (KBr) ν? _m ·. 3420 (br, OH), 1775 (β-lactam CO), 1695 '(-CO ₂ PNB) cm "

¹ H-NMR (dg-acetone) 6 : 9.44-7.72 (m, 8H, aromatic), 7.22-6.91 (m, 10H, diphenyl phosphate), 5.53, 5.27 (ABq , J = 14Hz, 2H, benzylic), 5.04 (t, J = 7.4Hz, 2H, N-CH ₂ ), 4.75 (s, 2H, CH ₂ OH) 4.5-3, 1 (m, 8H), 1.21 (d, J = 6.3 Hz, 2H, CHMe).

C. (5R, 6S) -3- [2- (3-hydroxymethylpyridinio) ethylthio] -6- [1- (R) -hydroxyethyl] -7-oxo-1-azabicyclo [3.2.0] hept-2 ene- 2-carboxvlate

CH ₂ OH

CO ₂ PNB

(0O) ₂ PO "

To a solution of p-nitrobenzyl- (5R, 6S) -3- [2- (3-hydroxymethylpyridinio) -ethylthio] -6- [1- (R) -hydroxyethyl] -7-oxo-1-azabicyclo [3.2. 0] hept-2-ene-2-carboxylate diphenyl phosphate (0.145 g, 0.194 mmol) in 10 ml of THF containing 5 drops of H 2 PO 4 is added to 6.0 ml of phosphate buffer (0.05 M, pH 7.4), 0.145 g of 10% palladium on carbon and 10 ml of ether. The mixture is hydrogenated (Parr) for 1 h at 40 psi, then filtered through a celite pad, the filter cake washed with a little H ₂ O and ether, the aqueous phase separated and extracted three times with ether. The aqueous solution is cooled to ⁰ ° C then .man _r and the pH with pH 7.4 buffer to 7.0. After removing the remaining volatiles in vacuo, the aqueous solution is applied to a C _1Q reverse phase column which is eluted with water. Lyophilization of the appropriate fractions gives the product (36 mg, 51%) as a light yellow solid. Further purification by reverse phase HPLC gives the pure product (31 mg, 41 %) as a solid. ,

IR (KBr)>? __ ": 3300 (br, OH), 1755 (β-lactam CO), 1590 '(-CO ₂ ") cm "'

¹ H-NMR (D ₂ O) (^: 8.78-7.94 (m, 4H, aromatic), 4.83 (t, J = 6.0 Hz, 2H, N-CH ₂ ), 4, 83 (s, 2H, CH ₂ OH), 4.16 (d of q, J = J '= 6.2 Hz, 1H, H-1 ^! ), 3.98 (d of t, J = 9.1 Hz, J '= 2.6 Hz, 1H, H-5), 3.75-3.20 (m, 3H), 3.20-2.65 (m, 2H), 1.22 (d, J = 6.4 Hz, 3H, CHMe)

UV (H ₂ O) X _max: 294 (£ 7614), 266 (£ 6936) nm T _1/2 (pH 7.4; 36.8 ⁰ C) 14.0 h.

Example 4

Preparation of (5R, 6S) -3- [2- (4-hydroxymethylpyridinio) ethylthio] -6- [1- (R) -hydroxyethyl] -7-oxo-1-azabicyclo- [5.2.01he-pt-2 -en-2-carboxylate

CH ₂ OH

A. 4-hydroxymethyl-1 - (2-mercaptoethyl) -pyridinium trifluoromethanesulfonate

! H ₂ OH + CF ₃ SO ₃ H + Γ \

HSCH ₂ CH ₂ N

CF ₃ SO ₃ "

CH ₂ OH

To a solution of 4-pyridinemethanol (1.635 g, 0.015 mol) in 10 mL of CH ₂ Cl ₂ is added dropwise under N ₂ at ⁰ ° C trifluoromethanesulfonic acid (1.327 mL, 0.015 mol). A yellow-brown oil separates quickly. An additional equivalent of ^ -pyridinemethanol (1.635 g, 0.015 mol) is then added to this mixture and the solvent removed under reduced pressure to give an oil. Ethylene sulfide (0.891 ml, 0.015 mol) is added to this oil and the resulting homogeneous mixture is heated to about 60 ^° C. for 3 hours on an oil bath. The reaction mixture is then taken up in 15 ml of water and the aqueous solution is washed with CH ₂ Cl ₂ (5 x 5 ml). After removing residual _% organic solvent in vacuo, the aqueous solution is placed on a C ^ g reverse phase column. It is eluted with water and the appropriate fractions are concentrated.

closing a to give an oil which was ₂ ° 5 ^vei in vacuo over P - ^ ^he dried to give the product (4.64 g, 97%) as a colorless oil.

IR (movie) \? _max : 3455 (st, OH), 2565 (schw, SH) cm ^{-1 1} H-NMR (dg-acetone) S : 9.07, 8.18 (ABq, J = 6.8 Hz, 4H, aromatic) , 5.03 (s, 2H, CH ₂ OH), 4.96 (1, J = 6.5Hz, 2H, N-CH ₂ ), 4.09 (br.s, 1H, -OH). 10

P-nitrobenzyl- (5R, 6S) -3- [2- (4-hydroxymethylpyridinio) ethylthio] -6- [i- (R) -hydroxyethylJ-7-0X0-1-azabicyclo3.2.0] hept-2 -en-2-carboxylate-diphenyl phosphate

CH ₂ OH j

0 CO ₂ PNB (0O) ₂ PO "j

To a solution of p-nitrobenzyl- (5R, 6S) -6- [i- (R) -hydroxyethyl] -3,7-dioxo-1-azabicyclo [3.2.0] heptane-2-carboxylate (0.348 g, 1 , 0 mmol) in 5 ml of dry acetonitrile are added dropwise under N ₂ at 0 ° C diisopropylethylamine (0.191 ml, 1.1 mmol) and then diphenylchlorophosphate (0.228 ml, 1.1 mmol). The resulting golden-yellow solution is stirred 40 min at ⁰ ° C, this solution is added to a solution of 4-hydroxymethyl-1- (2-mercaptoethyl) pyridinium trifluoromethanesulfonate (0.447 g,

1.4 mmol) in 1 mL acetonitrile, followed by diisopropylethylamine (0.191 mL, 1.1 mmol), with a reddish black, gummy product separating from the reaction mixture, after 20 min at ⁰ ° C, filter the reaction mixture and concentrate in vacuo. The residue is taken up in a small amount of acetonitrile-water (1: 1) and the mixture is placed on a C ^ Q reverse-phase column. Elute with 25% acetonitrile-water and then lyophilize the relevant fractions to give the product (0.353 g, 47%) as a cream solid

IR (KBr) \? _m ·. 3240 (br, OH), 1775 (β-lactam CO),

1695 (-CO ₂ PNB) cm ^{-1 1} H-NMR (.dg-acetone) <i: 9.24-7.84 (m, 8H, aromatic), 7.4-6.9 (m, 10H, Diphenyl phosphate), 5.52, 5.24 (ABq, J = 14Hz, 2H, benzylic), 5.15-4.80 (m, 4H), 4.45-3.05 (m, 7H), 1 , 35 (d, J = 6.6 Hz, 3H, CHMe).

C. (5R, 6S) -3- [2- (4-hydroxymethylpyridinio) ethylthio] -6- [1- (R) -hydroxyethyl] -7-oxo-1-azabicyclo [3.2.0] hept- 2 ene-2-carboxylate

OH

N / ^ Sy-CH ₂ OH

A mixture of p-nitrobenzyl- (5R, 6S) -3- [2- (4-hydroxymethylpyridinio) ethylthio] -6- [i - (R) -hydroxyethyl] -7-5 oxo-1-azabicyclo [3.2. 0] hept-2-ene-2-carboxylate diphenyl phosphate (0.348 g, 0.465 mmol) and 10% palladium on carbon (0.35 g) in 11 mL phosphate buffer (0.05 M, pH 7.4) » Hydrogenated 5 ml of THF and 10 ml of ether at 2.76 bar (40 psi) for 1.25 h. The mixture is filtered through a CeIite pad, the aqueous phase washed with ether (3x), the pH of the aqueous solution adjusted to 7.0 using another pH 7.4 buffer, residual volatiles removed in vacuo and puts the aqueous solution on a Cg reverse phase column. It is then eluted with 2% acetonitrile-water and then lyophilized to give a tan solid. This material is rechromatographed (Cj _Q reversed phase / H ₂ O) to give the desired product (0.060 g, 3696) as a light yellow solid / IR (KBr) C>: 3400 (br, OH), 1755 ( β-lactam OH), 1590 (-CO ₂ ") cm ^-1

¹ H-NMR (D ₂ O) cS ι 8.73, 7.96 (ABq, J = 6.8 Hz, 4H, aromat.), 4.93 (s, 2H, CH ₂ OH), 4.77 (t, J = 6.0 Hz, 2H, N-CH ₂ ), 4.15 (d of q, J = J '= 6.3 Hz, 1H, H-1 x), 3.96 (d of t, J = 9.2 Hz, J '= 2.6 Hz, 1H, H-5), 3.65-3.20 (m, 3H), 3.13-2.62 (m, 2H), 1.21 (d, J = 6.3 Hz, 3H, CHMe)

UV (H ₂ O) X _max : 295 (£ 6880), 256 (6 5595), 224 (£ 811i) nm t _Λ ΐ 2} (P ^H 7.4, 36.8 ° C) 14.5 h

 30

Example 5

Preparation of 3- [2- (i- (2-methylpyridinium)) ethylthio] 6a [1- (R) -hydroxyethyl] -7-oxo-1-azabicyclo [3.2.0] hept- 2-ene-2 carboxylate

COO

A. 1- (2-Mercaptoethyl) -2-methylpyridinium methanesulfonate

zf \

4

 P:

MsOH

55 ⁰ C,

21 h

MsO

To a suspension of 2-methylpyridinium methanesulfonate in 2-methylpyridine prepared by adding methanesulfonic acid (0.65 ml, 0.010 mol) to cold 2-methylpyridine (2.17 ml, 0.022 mol) is added ethylene sulfide (0.655 ml, 0.011 mol), the reaction mixture is stirred for 21 h under a nitrogen atmosphere at 55 ⁰ C, cooled to 23 ° C and diluted with 5 ml of water. The aqueous solution is washed with ether (6x4 ml). It removes traces of organic solvents by pumping. The solution is then placed on a column (2.5 x 10.0 cm) of μ-Bondapack C-18, the column is eluted with water and lyophilized the appropriate fractions, giving 2.13 g (85%) of the title compound receives.

IR (film) v> - 2520 (SH), 1623 (pyridinium), 1574, 1512, 1485, 1412, 1195 (sulfonate), 1038 cm ^"1

¹ H-NMR (DMSO-dg + D ₂ O) O: 2.37 (3H, s, CH ₃ SO ₃ "), 2.83 (3H, s, CH ₃ on pyridinium), 3.09 (2H, J = 6.9 Hz, CH ₂ S), 4.71 (2H, t, J = 6.9 Hz, CH ₂ N ⁺ ), 7.93 (2H, m, Hm of pyridinium), 8.44 ( 1H, m, Hp of pyridinium), 8.89 (1H, m, Ho of pyridinium) _max : 266 (3550) _{m /} u.

10 UV (H ₂ O) A

For example, p-nitrobenzyl-3- [2- (1- (2-methylpyridinium) ethylthioJ-α-ti-1-hydroxyethyl ] -1-oxo-i-azabicyclo [3.2.0] hept-2-ene-2-carboxylate -dipheny phosphate!

1) NEt (IPr) ₂

2) ClP (OPh) ₂

3) HS COOPNB

0 'COOPNB (PhO) ₂ PO "

(To a cold ⁽⁰ ° C) solution of p-nitrobenzyl 6 <x [i-5 (R) -hydroxyethyl] -3,7-dioxo-1-azabicyclo [3.2.0 Jheptan-2-carboxylate 0.523 g , 1.50 mmol) in acetonitrile (6 ml) (stored under nitrogen) are added diisopropylethylamine (0.314 ml, 1.80 mmol) followed by diphenylchlorophosphate (0.373 ml, 1.80 mmol). The reaction mixture was stirred for 30 min at ⁰ ° C and treated with a solution of 1- (2-mercaptoethyl) -2-methylpyridiniummethansulfonat (0.530 g, 2.16 mmol) in 18 ml of acetonitrile and then with diisopropylethylamine (0.314 ml, 1, 8 mmol). The reaction mixture is stirred for 1 h at ^{0 0} C, diluted with cold (O ⁰ C) water (26 ml) and gives the mi

on a U-Bondapak C ^ g column (3.5 χ 7.0 cm). The column is eluted with 25% acetonitrile-75% water and 50% acetone tril-50% water. After lyophilization of the appropriate fractions, 1.06 g (96%) of the title compound are obtained as a yellowish powder.

IR (KBr) v> _max : 3650-3100 (OH), 1770 (C = O of β-lactam), 1695 and 1690 (C = O of the PNB ester), 1630 (pyridinium), 1595 (phenyl), 1518 (NO ₂ ), 1335 (NO ₂ ), 890 cm ^-1 (NO ₂ )

¹ H-NMR (DMSO-dg) S: 1.15 (3H, d, J = 6.1 Hz, CH ₃ CHOH), 2.87 (s, _CH3 on pyridinium), 3.6-4.4 (2H, m, H-5, CH ₃ CHOH), 4.75 (2H, m, CH ₂ N ^+), 5.37 (center of ABq, J = 14 Hz, _CH2 of PNB), 6.5 -7.4 (1OH, m, phenyl), 7.70 (2H, d, J = 8.8Hz, Ho of PNB), 8.0 (2H, m, Hm of pyridinium), 8.24 (2H , d, J = 8.8 Hz, Hm of PNB), 8.50 (1H, m, Hp of pyridinium), 8.95 (1H, br.d, J = 6.1 Hz, Ho of pyridinium)

UV (H ₂ O) A _max : 265 (£ 11,990), 314 (£ 8020) iyi.

C. 3- [2- (1- (2-Methylpyridinium)) ethylthio] -6a [i (R) -hydroxyethyl] -7-oxo-1-azabicyclo [3.2.0] hept-2 en-2- carboxylate

OH

10% Pd / C, H _{2 v}

THF, ether, buffer

COOPNB

(PhO) ₂ PO "

To a solution of p-nitrobenzyl-3- [2- (i- (2-methylpyridinium)) ethylthio] -6a- [i- (R) -hydroxyethyl] -7-5 oxo-1-azabicyclo [3.2.0 ] Hept-2-ene-2-carboxylate diphenyl phosphate (0.66 g, 0.90 mmol) in wet tetrahydrofuran (34 mL) is added ether (34 mL), potassium phosphate monobasic sodium hydroxide buffer (0 → 15 M, 16, 5 ml, pH 7.22) and 10% palladium on carbon (0.66 g) and hydrogenates the resulting mixture for 1.25 h at 1.76 bar (40 psi) pressure and 23 ° C. The organic layer is separated off and extracted with buffer (2 × 6 ml). The layers of water are combined, filtered through a pad of celite, washed with ether (40 ml), traces of organic solvents are removed by applying a vacuum and applied to a / U Bondapak Cjg column (2.5 x 10 cm). The column is eluted with water and the appropriate fractions are lyophilized to give 0.098 g (31%) of the title compound as a yellowish powder. IR (KBr) C>: 3650-3100 (OH), 1755 (C = O of β-lactam),

HLcLX. a ~

1630 (pyridinium), 1595 cm ^-1 (carboxylate) ¹ H-NMR (D ₂ O) <S: 1.20 (3H, d, J = 6.3 Hz, CH ₃ CHOH), 2.83 (s, _CH3 on pyridinium), 2.7-3.1 (5H, H-4, _CH3 on pyridinium), 3.1-3.7 (3H, m, CH ₂ S, H-6), 3.90 (dd, J = 9.1 Hz, J = 2.6 Hz, H-5), 3.1 (m,

CH ₃ CHOH), 4.78 (t, J = 6.2 Hz, CH ₂ N ⁺ ), 7.8 (2H, m, .Hm of pyridinium), 8.3 (1H, m, Hp of pyridinium) , 8.65 (1H, s, Ho of pyridinium) UV (H ₂ O) * _max : 268 (19350), 196 (£ 8840) nyu [a] J p = + 41 °

^ 1/2 ⁼ h ^ 5.0 (measured at a concentration of 10 ~ ⁴ M in phosphate buffer pH 7.4 at 36.8 ⁰ C) *

Example 6

Preparation of 3- [2- (1- (4-methylpyridinium)) ethylthio] -6a- [1- (R) -hydroxyethyl] -7-oxo-1-azabicyclo [3.2.0] hept-2-ene -2-carboxylate

COO j

A. 1- (2-Mercaptoethyl) -4-methylpyridinium methanesulfonate

. + MsOH

55 ° C, 24 h

MsO '

To a suspension of 4-picolinium methanesulfonate in 4-picoline prepared by adding methanesulonic acid (0.65 ml, 0.010 mol) to 4-picoline (2.14 ml, 0.022 mol) is added under cooling ethylene sulfide (0.655 ml, 0.011 Mole), the reaction mixture is stirred for 24 hours under a nitrogen atmosphere at 55 ° C, cooled to 23 ° C and diluted with water (5 ml) and ether (10 ml). The organic layer is separated, the aqueous layer washed with ether (5x5 ml) and added to a 2.5 × 10 cm column of U-Bondapak C ^ g after removing traces of ether under reduced pressure. Elute the column with a 15% acetone / tri1-85% water mixture and lyophilize the appropriate fractions. This gives 2.66 g (100%) of a colorless syrup.

IR (film) ^ _max : 2500 (SH) / 1640 (pyridinium), 1572, 1520, 1478, 1200 (sulfonate), 1040, 833 and 768 cm ^-1

¹ H-MR (DMSO-dg) 6: 2.31 (3H, s, CH ₃ SO ₃ "), 2.62 (s, _CH3 on pyridinium), 2.2-2.9 (4H, SH, CH ₃ on pyridinium), 3.04 (2H, m, CH ₂ S), 4.68 (2H, t, J = 6.4Hz, CH ₂ N ⁺ ), 8.01 (2H, d, J = 6.6 Hz, Hm of pyridinium), 8.89 (2H, d, J = 6.6 Hz, Ho of pyridinium), UV (H ₂ O) A _max : 256 (64100), 221 (£ 7544) myu.

B. 1 - (2-Mercaptoethyl) -4-methylpyridinium p-toluenesulfonate

TsO "

To a suspension of p-toluenesulfonic acid (1.72 g,

0.01 mole) in benzene (6.5 ml) is added 4-picoline (1.17 ml, 0.012 mole), the resulting mixture is stirred for 30 minutes under a nitrogen atmosphere at 23 ° C, treated with ethylene sulfide (0.65 ml, 0.011 mol) and stirred for 24 h at 75 ⁰ C is added to more ethylene sulfide (0.65 ml, 0.011 mol), stirred for a further 24 h at 75 ⁰ C, the reaction mixture is cooled to 23 ° C and diluted with 5 ml of water and 8 ml of ether, the aqueous layer is separated off, washed with ether (3 × 8 ml), traces of organic solvents are removed.

Medium in vacuo and chromatographed on / U-Bondapak

C-18 using water as eluent. This gives 2.94 g (90%) of the title compound as a colorless syrup

IR (movie) v>'. 2510 (SH), 1640 (pyridinium), 1595, 1582, 1475, 1475, 1200 (sulfonate), IO3I, 1010, 818 cm ^-1

¹ H-NMR (DMSO-dg) 6 : 2.29 (3H, s, CH, on pyridinium), 2.61 (s, CH, Ph), 2.4-2.8 (4H, SH, CH ^ Ph), 3.03 (2H, m [addition of D ₂ O gives a t, J =

6.4 Hz, at 3.04], CH ₂ S), 4.68 (2H, t, J = 6.4 Hz, CH ₂ N), 7.11, 7.49 (4H, 2d, J = 7.9 Hz, phenyl), 8.00 (2H, d, J = 6.5 Hz, Hm of pyridinium), 8.89 (2H, d, J = 6.5 Hz, Ho of pyridinium) UV (H ₂ O) \ _max : 256 (^ 4315), 222 (£ 17045) myu.

C. p-Nitrobenzyl-3- [2- (1- (4-methylpyridinium) -ethylthio] -6a [1- (R) -hydroxyethyl] -7-oxo-1-azabicyclo- Γ3.2.0] hept-2-ene-2-carboxylate dipheny! phosphate

1) NEt (iPr) ₂

2) ClP (OPh) ₂

3) HS

4) NEt (IPr) ₂

COOPNB

(PhO) -PO COOPNB ² "0

(To a cold ⁽⁰ ° C) solution of p-nitrobenzyl 6oc- [i- (R) -hydroxyethyl] -3,7-dioxo-1-azabicyclo [3.2.0 Jheptan-2-carboxylate 0.522 g ,, 1 5 mmol) in acetonitrile (6 ml) (stored under nitrogen) are added diisopropylethylamine (0.314 ml, 1.8 ml of oil) followed by diphenylchlorophosphate (0.373 ml, 1.9 mmol), the reaction mixture is stirred for 45 min and treated added dropwise with a solution of 1- (2-mercaptoethyl) -4-methylpyridinium methanesulfonate (0.539 g, 2.16 mmol) in acetonitrile (1.8 mL) followed by diisopropylethylamine (0.314 mL, 1.8 mmol). The reaction mixture is stirred

1 h at ⁰ ° C, diluted with cold (0 ⁰ C) water (24 ml), and outputs a / u-Bondapak C-18 column (2.5 x 8.5 cm). The column is eluted first with a 25% acetonitrile-75% water mixture (100 ml), then with a 50% acetonitrile-50% water mixture (100 ml), and after lyophilization of the appropriate fractions 0.91 g (83 %) of the title compound as a yellowish powder. IR (KBr) Q _m ·. 3700-2800 (OH), 1770 (C = O of β-lactam), 1700 (C = O of the PNB ester), 1640 (pyridinium), 1595 (phenyl), 1520 (NO ₂ ), 1340 (NO ₂ ) , 890 cm " ¹ (NO ₂ )

¹ H-NMR (DMSO-d ₆ ) 6 : 1.16 (3H, d, J = 6.2 Hz, CH ₃ CHOH), 2.61 (s, CH ₃ am. Pyridinium), 3.1-3 , 7 (3H, m, H-6, _CH2 S), 3.7-4.4 (2H, m, H-5, CH ₃ CHOH), 4.79 (2H, br t, J = 6 , 3 Hz, CH ₂ N ⁺ ), 5.17 (d, J = 4.9 Hz, OH), 5.37 (center of ABq, J = 14.1 Hz, CH ₂ of PNB), 6.7 -7.4 (1OH, m, phenyl), 7.69 (2H, d, J = 8.8 Hz, Ho of PNB), 8.00 (2H, d,

J = 6.5 Hz, Hm of pyridinium), 8.23 (2H, d, J = 8.8 Hz, Hm of PNB), 8.92 (2H, d, J = 6.5 Hz, Ho of pyridinium )

UV (H ₂ O) A _max : 262 (ε 10835), 311 (8: 9670) iyi Analysis: Calculated for C ₃₆ H ₃₆ N ₃ O _1Q SP.1.5H ₂ O: C, 56.84% H 5, 17% N 5.52% S 4.21% Found: 56.89 5.13 5.19 4.41.

D. 3- [2- (1- (4-Methylpyridinium)) ethylthio] -6a [i- (R) -hydroxyethyl] -7-oxo-1-azabicyclo [3.2.0] hept-2-ene 2- carboxylate_

OH

Il _

COOPNB (PhO) ₂ PO

10% Pd / C, Η _2χ

THF, ether, buffer

COO "

To a solution of p-nitrobenzyl-3- [2- (1- (4-methylpyridinium) -ethylthio] -6a- [i- (R) -hydroxyethyl] -7-oxo-1-azabicyclo [3.2.0] Hept-2-ene-2-carboxylate diphenyl phosphate (0.587 g, 0.80 mmol) in wet tetrahydrofuran (30 mL) is added with ether (30 mL), potassium phosphate monobasic sodium hydroxide buffer (0.15 M, 14.7 mL, pH 7.22) and 10% palladium on carbon (0.59 g), the resulting mixture hydrogenates for 1.25 h at 23 ° C under a pressure of 1.76 bar (40 psi), separating the organic layer and extracted with the buffer (2 × 6 ml). The aqueous extracts are combined, filtered through a pad of celite, washed with ether (3 × 20 ml), traces of organic solvents are removed by pumping and applied to a / U Bondapak C-18 column (2.5 cm × 10 cm). , Elution of the column with water and lyophilization of the appropriate fractions gives 0.13.6 g (49%) of the title compound as a yellowish powder.

IR (KBr) O

Max

3700-3000 (OH), 1770 (C = 0 of β-lactam),

-1

1642 (pyridinium), 1592 cm "'(carboxylate) ₁ H-NMR (D ₂ O) 6: 1.19 (3H, t, J = 6.3 Hz, CH ₃ CHOH), 2.59 (3H, s, CH ₃ on pyridinium), 2.84 (d, J = 9.1 Hz,

H-4), 2.90 (d, J = 9.1Hz, H-4), 3.0-3.6 (3H, m, CH ₂ S, H-6), 3.86 (dd, J = 9.1 Hz, J = 2.6 Hz, H-5), 4.12 (m, CH ₃ CHOH), 4.5-4.9 (CH ₂ N ⁺ masked

by HOD), 7.80 (2H, d, J = 6.6 Hz, Hm of pyridinium), 8.58 (2H, d, J = 6.6 Hz, Ho of pyridinium), UV (H ₂ O) ? \ _max : 256 (£ 5510), 262 (£ 5360), 296 (6 7O5O) nm [cc] Jp = + 20.8 ° (C 0.48, H ₂ Oj

Ty ₂ = 12,8h (measured at a concentration of

10 "^ M in a phosphate buffer pH 7.4 at 36.8 ⁰ C),

Example 7

Preparation of (5R) -3- [2- (4-methylthiopyridinio) ethylthio] - (6S) - [(1R) -hydroxyethyl] -7-oxo-1-azabicyclo [3.2.O] -hept -2-ene -2-carboxylate

A. 4-methylthiopyridine

/ ^ \ 1) MeJ / EtOH

2) NaOH

4-Mercaptopyridine (5.55 g, 50.0 mmol, Aldrich) is dissolved in 50 ml of boiling, absolute ethanol. The insoluble material is filtered off over CeIite. The filtrate was heated to re-releasing and when it is cooled to about 50 ⁰ C, is added methyl iodide (3.17 ml, 51.0 mmol; Aldrich) at a time to · The mixture is cooled to crystallize. , The solid is filtered off,

to obtain 6.77 g (26.7 mmol, yield 53.5%) of the title compound as a hydroiodide.

¹ H-NMR (D ₂ O) S : 2.70 (3H, s, -SCH ₃ ) and 7.65-7.77-8.35-8.48 ppm (4h, A ₂ B ₂ type, aromat.Hs)

IR (Nujol) \? _v : 1615, 1585 (aromat.) and 780 cm " ¹ max r

UV (H ₉ O) λ _m z 227 (L 2.02 χ 10 ^) and 298 nm (£ 1.64 x1

The hydroiodide (6.33 g »25.0 mmol) is dissolved in 40 ml of water, insoluble material is removed, washed with 10 ml of water, added to the filtrate at 0-5 ° NaOH pellets (5 g) and extracted with Et ₂ O (3 x 25 ml), the aqueous layer being saturated with NaCl, the combined organic extracts washed with brine (2 χ), dried (MgSO 4) and concentrated to give 2.92 g (23.4 mmol , Total yield 50%) of the title compound as an oil. ¹ H-NMR (CDCl ₃₎ £: 2.48 (3H, s, -SCH ₃₎ and 7,03-7,13-8,38-8,48 ppm (4H, A ₂ B ₂ ~ type aromat, .Hs) IR (film) wL _ev : 1580 and 800 cm ^-1 . The preparation of this compound is described by King and Ware in J. Chem. Soc., 873 (1939) The process described therein was used.

For example, 4-methylthio-N- (2-mercaptoethyl) pyridinium methanesulfonate

SMe

50-60

CH ₃ SO ₃

4-Methylthiopyridine (2.75 g, 22.0 mmol) is added slowly, while cooling in an ice bath, to methanesulfonic acid " ¹ " (0.65 ml, 10.5 mmol). To this solid is added ethylene sulfide ⁺ (0.66 ml, 11.0 mmol), Aldrich) and the mixture is heated 21 h at 50 to 60 ⁰ C. With the progress of reaction, the solid goes into solution. After cooling, the reaction mixture is dissolved in 5 ml of water, washed with Et ₂ O (5 × 4 ml), the flaky aqueous layer is filtered through Celite, and the filtrate is purified by reverse phase silica gel column chromatography (C ^ micro Bondapack 10 g). , eluting with water. Fractions of 10 ml each were collected. . The fractions 2 and 3 combined one and cleans it in turn ^m i "t ^ of the reverse phase column Fraction 2 yields 1.258 g (4.48 mmol, yield 42.6%) of the title compound as a viscous oil.

, ¹ H-NMR (DMSO-d ₆ , CFT-20) £: 2.32 (3H, s, MeSO ₃ "), 2.72 (3H, s, -SMe), 2.68 (1H, m, SH ), 2.9-3.2 (3H, m, -CH ₂ S-), 4.59 (2H, t, J = 6.4 Hz, -CH ₂ N ⁺ ), 7.97 (2H, d , J = 7.2 Hz, aromat.Hs) and 8.72 ppm (2H,

d, J = 7.2 Hz, aromat.Hs) IR (neat) v> _v : 1630, 1200 (br., -SO, ""), 785 and 770 cm ^-1

⁺ These reagents were distilled before use

 25

C. (5R) -p-nitrobenzyl-3- [2- (4-methylthiopyridino) -ethyl]

Γ3.2.01hept-2-en-2-carboxylate chloride 30

I) ClPO ((V ').

CO ₂ PNB

_Ms0 -

OH

SMe

CO ₂ PNB

20 25 30 35

To a solution of (5R) -p-nitrobenzyl-3,7-dioxo (6S) - [(1R) -hydroxyethyl] -1-azabicyclo [3.2.0] heptane (2R) -carboxylate (475 mg , 1.36 mmol) and diisopropylethylamine (0.24 ml, 1.4 mmol) in 5 ml of CH ₃ CN are added under nitrogen atmosphere at 0 to 5 ⁰ C diphenylchlorophosphate (0.29 ml, 1.41 mmol), stirred the mixture for 3o min at 0 to 5 ° C, is added to this mixture, an oily suspension of 4-methylthio-N- (2-mercaptoethyl) pyridinium methanesulfonate (678 mg, 1.45 mmol, 60% purity) in 1 , 5 ml of CHUCN and then diisopropylethylamine (0.24 ml, 1.4 mmol) and the mixture is stirred for 1 h at 0 to 5 ° C Immediately after the addition of the base, a yellow precipitate forms. The precipitate is filtered off and washed with 3 ml of cold CH 2 CN to give 413 mg of a yellowish solid. This is triturated with 10% MeOH in water (5 ml) to give 341 mg (0.618 mmol, yield 45.4%) of the title compound as white crystals; Mp 118 to 120 ^° C.

¹ H-NMR (DMSO-dg, CFT-20) S ; 1.16 (3H, d, J = 6.1 Hz, 1'-CH ₃ ), 2.72 (3H, s, -SCH ₃ )

3.1-3.7 (5H, m),

3.7-4.3 (2H, m), 4.71 (2H, t, J = 6.3Hz, -CH ₂ N ⁺ ), 5.15 (1H, d, J = 4.9Hz, OH), 5.20-5.35-5.40-5.55

(2H, ABq, CO ₂ CH ₂ -Ar), 7.70 (2H, d, J = 8.8 Hz, nitrophenyl Hs), 7.97 (2H, d, J = 7.0 Hz, pyridinio Hs), 8.25 (2H, d, J = 8.8 Hz, nitrophenyl Hs) and 8.76 ppm (2H, d, J = 7.1 Hz, pyridinio-Hs) IR. (NuJoI) \? _ _n __: 3250 (OH), 1775 (β-lactam), 1700 (ester)

and 1625 cm (pyridinio) UV (abs.EtQH) \ _max : 308nm (£ 4.47 x 10 ⁴ ) [a] jp = + 24.8 ° (c 0.5, MeOH)

Analysis: for C _2Zf H ₂ gN, OgS ₂ Cl.H ₂ O.

Calculated: C 50.56% H 4.95% N 7.37% Found: 50.63 4.72 6.89.

D. (5R) -3- [2- (4-Methylthiopyridinio) -ethylthio] - (6S) - [(1R) -hydroxyethyl] -7-oxo-1-azabicyclo [3.2.0] hept-2- ene 2-carboxylate

H ₅ / Pd-C

SMe _ £ ___

PH 7.4 buffer

CO ₂ PNB

(5R) -p-nitrobenzyl-3- [2- (4-methylthiopyridinio) ethylthio] - (6S) - [(1R) -hydroxyethyl] -7-oxo-1-azabicyclo [3.2.0] hept-2 -en-2-carboxylate chloride (380 mg, 0.688 mmol) is dissolved in 31.5 ml of THF and a pH 7.40 phosphate buffer (31.5 ml, 0.05M, Fisher) diluted with Et ₂ O (31 , 5 ml), this solution is mixed with 10% Pd-C (380 mg, Engelhard) and

hydrogenated at room temperature for 1 h with a Parr shaker at 2.4 bar (35 psi). The aqueous layer is filtered through Celite to remove the catalyst and the celite pad washed with water (2x5 mL). The filtrate and washings are combined and washed with Et ₂ O (2 × 30 ml) The aqueous layer is freed from organic solvents under reduced pressure and purified by reverse phase column chromatography (CJG-Microbondapak, 13 g, Waters Associates). eluting with water Fractions with a UV absorption at 307 nm are collected (about 11) and lyophilized to give 127 mg (0.334 mmol, yield 48.5%) of the title compound as a yellowish powder.

¹ H-NMR (D ₂ O, CFT-20) S ¹ 1.20 (3H, d, J = 6.4 Hz, 1'-CH ₃ ), 2.64 (3H, s, -SCH,), 2.81 (2H, m, -SCH ₂ -), 3.19 (1H, dd, J _6-1 I = 6.1 Hz, Jg_ ₅ = 2.6 Hz, 6-H), 3.32 ( 2H, dd, J = 11 Hz, J = 5.5 Hz, 4-Hs), 3.92 (1H, dt, J = 9.2 Hz, JV ₆ = 2.6 Hz, 5-H), 4 , 1 (1H, m, 1'-H), 4.61 (2H, t, J = 5.9 Hz, -CH ₂ N ⁺ ), 7.70 (2H, d, J = 7.1 Hz, aromat.Hs) and 8.40 ppm (2H, d, J = 7.1 Hz, aromat.Hs)

IR (KBr, Pill) <? _m ·. 3400 (OH), 1750 (.beta.-lactam), 1630 (pyridinium) and 1590 cm-(carboxylate) UV (H ₂ O) X _max: 231 (£ 9800) and 307 nm (£ 25,000) Ο] ^{2, 3,} m + 3.14 ° (c 0.5, H ₂ O).

Example 8

Preparation of 3- [2- (3-methoxy-1-pyridinium) -ethylthio] 6a- [i ^f - (R) -hydroxyethyl] -7-oxc-1-azabicyclo [3.2.0] hept- 2-ene 2-carboxylate

OMe 35

A. 1- (2-Mercaptoethyl) -3-methoxypyridinium methanesulfonate

nat.

_q MsOH MeO

To pre-cooled (5 ⁰ C) 3-methoxypyridine (698 mg, 6.4 mmol) are added dropwise methanesulfonic acid (0.216 ml, 3.05 mmol) and ethylene sulfide (0.19 ml, 3.2 mmol). The mixture is then heated for 18 h at 60 ⁰ C, cooled to 20 ⁰ C, diluted with water (10 ml) and washed with ether (3 x 10 ml). The aqueous phase is treated at high vacuum for 15 minutes and added to a C 2 -C 4 reverse phase column. The title compound is eluted with water. Combine the appropriate fractions and concentrate under high vacuum to give the desired thiol (61.6 mg, 76.3% yield).

IR (CH ₂ Cl ₂ ) ^ _11121x : 2550 (schw, SH) and 1620, 1600,

1585 cm ^-1 "(m, aromat.) ¹ H-NMR (DMSO-dg) S ; 8.90-7.90 (4H, m, aromat.CH), 4.72

(2H, t, J = 6.6Hz, CH ₂ N ⁺ ), 4.01 (CH, s, OCH ₃ ), 3.5-3.0 (m, hidden. CH ₂ S), 2.66 (1H, dd, J = 9.5 Hz, J = 7.5 Hz, SH) and 2.31 ppm (3H, s, CH ₃ SO ₃ -).

For example, p-nitrobenzyl-3- [2- (3-methoxy-1-pyridinium chloride) -ethylthio] -6oc- [i '- (R) -hydroxyethyl] -7-oxo-1-azabi-

cvclo Γ3.2.01 hept-2-ene-2-carboxylate

^C1 OMe,

4) EtNC-O ₂

CO ₂ PNB CO ₂ PNB

A cold ⁽⁰ ° C) solution of p-nitrobenzyl 6a- [i '- (R) -hydroxyethyl] -3,7-dioxo-1-azabicyclo [3.2.0] heptane-2-carboxylate (1.04 g , 3 mmol) in 12 ml of acetonitrile are treated dropwise with diisopropylethylamine (0.63 ml, 3.6 mmol) and diphenylchlorophosphate (0.75 ml, 3.6 mmol) and stirred for 30 min at ⁰ ° C. The resulting enol phosphate is treated with 1- (2-mercaptoethyl) -3-methoxypyridinium methanesulfonate (1.14 g, 4.30 mmol) in 7 mL of CH 2 CN, 0.63 mL (4.30 mmol) of diisopropylethylamine, stirred for 30 min and cooled 30 min at -10 ⁰ C. the precipitated solid was filtered from the mixture, washed with cold acetonitrile (2 ml) and dried to afford the title compound is obtained (1.32 g, yield 82%).

IR (Nujol) J "'. 3320 (m, OH), 1780, 1765 (St., .Beta.-lactam

C = O), 1700, 1695 (m, ester C = O) and 1520 cm ^-1 (st, NO ₂ ) ¹ H-NMR (DMSO-dg) 6 : 9.01 (1H, bs, H-3 aromat .), 8,75

(1H, bd, J = 5.4Hz, H-6 aromat.), 8.35-7.95 (4H, m, H-aromat.), 7.70 (2H, d, J = 7.7 Hz, H-aromat.), 5.37 (2H, center of ABq, J = 13 Hz, _CH2 PNB), 5.17 (1H, d, J = 4.9 Hz, OH), 4.87 ( 2H, t, J = 6.3Hz, CH ₂ -N ⁺ ), 4.35-3.75 (2H, m, H-5 and H-1 x), 4.00 (3H, s, OCH, ), 3.56 (part of t, J = 6.3 Hz, CH ₂ S), 3.5-3.20 (3H, m, H-6, H-3) and 1.16 ppm (3H, d, J = 6.1 Hz, _CH3 CHO).

C. 3- [2- (3-Methoxy-1-pyridinium) ethylthio] -6 <x- [i '- (R) -hydroxyethyl] -7-oxo-1-azabicyclo [3.2.0] hept-2 -en-2 car boxy lat · _:

Cl "OMe

CO ₂ PNB

A solution of p-nitrobenzyl-3- [2- (3-methoxy-1-pyridinium chloride) ethylthio] -6a [i * - (R) -hydroxyethyl] -7-oxo-1-azabicyclo [3.2.0] hept-2-ene-2-carboxylate (600mg, 1.12mmol) in THF (25ml), ether (25ml) and pH 7.4 phosphate buffer (0.1M, 25ml) are hydrogenated for 1h a Parr shaker at 2.76 bar (40 psi) above 10% Pd / C. Dilute the mixture with ether and filter the aqueous phase through a # 52 hardened filter paper. The aqueous layer is washed with ether (2 × 20 ml), a vacuum is applied and the mixture is placed on a silica gel reverse phase column. The title compound is eluted with water containing 2% and 5% acetonitrile. The appropriate fractions are combined and lyophilized to give a yellow solid which is purified by HPLC to give the penem carboxylate (150 mg, IR (Nuool).

Max

1750 (st, β-lactam C = O) and 1580 cm

(st, carboxylate)

¹ H-NMR (D ₂ O) S : 8.55-8.30 (2H, m, H-2, H-6 aromat.), 8.17-7.75 (2H, m, H-3, H-4 aromat.), 4.77 (2H, t, J = 5.9 Hz, CH ₂ N ⁺ ), 4.10 (1H, part of 5 lines, J = 6.3 Hz, H-1 ' ), 3.97 (3H, s, OCH ₃ ), 3.85, 3.82 (2 lines, part of dt, J = 2.6 Hz, part of H-5), 3.42 (2H, t , J = 5.9 Hz, CH ₂ -S), 3.25 (1H, dd, J = 6.1 Hz, J = 2.6 Hz, H-6), 2.99-2.60 (2H , 6 lines, part of H-4) and 1.20 ppm (3H, d, J = 6.4 Hz, CH ₃ )

UV (H ₂ O, c 0.05) λ _{Ώ & χ} : 290 (£ 10517), 223 (β 6643) T ₁ ^ ₂ (0.1 M pH 7.4 phosphate buffer, 37 ^° C) = 20 h.

15 At play

Preparation of (5R, 6S) -3- [2- (3-methylthiopyridinio) ethylthio] -6 - [i -] - hydroxyethylj ^ oxo-i-azabicyclo [3.2.Q1hept-2-ene-2-carboxylate

A. 5-Methylthio-i- (2-mercai-3-ethyl) -pyridinium chloride

SMe

SMe

+ HCl +

4 HSCH ₂ CH

'• Ü

To a solution of 3-methylthiopyridine (2.00 g, 0.016 mol) (prepared by the method of JAZoltewiez and C. Nisi, J. Org. Chem., 4, 765, 1969) in 10 ml of ether is added 15 ml 1N HCl, shake the mixture thoroughly,

separates the aqueous phase, washed with 10 ml of ether and then concentrated. The remaining hydrochloride is then dried in vacuo (PpO ₁ -) to give a white solid. To this hydrochloride solid is added 3-methylthiopyridine (1.88 g, 0.015 mol) and ethylene sulfide (0.89 mL, 0.015 mol) and the resulting mixture (oil bath) heated at 55-65 ° C under N ₂ for 15 h. This gives a slightly cloudy oil which is taken up in 125 ml of water and washed with CHpCl ₂ . The aqueous solution is concentrated to about 25 ml, then a few drops of acetonitrile are added to make the mixture homogeneous, the resulting aqueous solution is applied to a C ^ g reverse phase column, eluted with water, and then the relevant fractions are concentrated the product is obtained as a pale yellow, viscous oil (2.66 g, 80%). IR (FiImW _ma i 2410 (br, -SH) cm ^{"1 1} H-NMR (dg-DMSO + DPO) 6: 8.88 to 7.88 (m, 4H, aromatic), 4.70 (t,. J = 6.5 Hz, 2H, N-CH ₂ ), 3.08 (oblique t, J = 6.5 Hz, 2H, S-CH ₂ ), 2.64 (s, 3H, S-Me).

P-nitrobenzyl- (5R, 6S) -3- [2- (3-methylthiopyridinio) -ethylthio] -6 - [1 - (R) -hydroxyethyl] -7-oxo-1-azabicyclo ["5.2. 0 ~ lhept-2-en-2-carboxvlat chloride

CO ₂ PNB

^ \- ".s / ~ \ _ri

CO ₂ PNB

A solution of p-nitrobenzyl- (5R, 6S) -6- [i- (R) -hydroxyethyl] -3,7-dioxo-1-azabicyclo [3.2.0] heptane-2-carboxylate (0.522 g, 1, 50 mmol) in 7 ml of dry acetonitrile is cooled to ⁰ ° C., then diisopropylethylamine (0.287 ml, 1.65 mmol) is added dropwise, diphenylchlorophosphate (0.322 ml, 1.65 mmol) is added dropwise to the resulting tan solution the reaction mixture is then added at ⁰ ° C. for 30 minutes. Diisopropylethylamine (0.313 ml, 1.80 mmol) is then added followed by a solution of 3-methylthio-1- (2-mercaptoethyl) -pyridinium chloride (0.398 g, 1.80 mmol). in 0.70 ml of dry DMF. About 1 minute after completion of the addition, a precipitate separates from the reaction mixture. Further cooling to -10 ° C for 10 minutes gives a solid, orange mass. This solid is then triturated with acetonitrile and the residue is filtered off. The residue is washed with acetonitrile, then acetone and finally dried in vacuo to give the product (0.455 g, 55%) as a cream solid. The combined filtrate is concentrated to give a yellow oil, which is taken up in as little as possible acetonitrile and cooled to 0 ° C for 30 min. This mixture is filtered, giving another 0.139 g of the product as a pale yellow solid. The total yield is 0.594 g (72%). IR (KBr) v> _ _v 3345 (br, -OH), 1770 (.beta.-lactam CO), 1680 (CO ₂ PNB) cm "'

¹ H NMR (d ₆ -DMSO) S : 8.98-7.96 (m, 4H, pyridinium aromat.). 8.20-7.65 (ABq, J = 7.0 Hz, 4H, PNB aromat.) 5.53-4.80 (m, 4H), 4.3-3.7 (m, 2H), 3 , 6-3.25 (m, 6H), 2.66 (s, 3H, S-Me), 1.16 (d, J = 6.0 Hz, 3H, CHMe).

C. (5R, 6S) -3- [2- (3-Methylthiopyridinio) -ethylthio] -6- [1- (R) -hydroxyethyl] -7-oxo-1-azabicyclo [3.2.0 Jhept- 2-ene -2-carboxylate

SMe

CO ₂ PNB

To a mixture of p-nitrobenzyl- (5R, 6S) -3- [2- (3-methylthiopyridinio) -ethylthlo] -6- [1- (R) -hydroxyethyl] -7-oxo-1-azabicyclo [3.2. 0] -hept-2-ene-2-carboxylate chloride (0.551g, 1.0mmol) and 10% palladium on carbon (0.55g) in 25ml phosphate buffer (0.05M, pH 7, 4) are added 5 ml of THF and 25 ml of ether. The mixture is hydrogenated (Parr) for 1 h at 2.76 bar (40 psi). The reaction mixture is then filtered through Celite and the filter cake is washed with water and ether. The aqueous phase is separated off, washed with additional ether (3x), residual organic solvents are removed in vacuo and the aqueous solution is cooled to ⁰ ° C. and the pH is adjusted to 7.0 with saturated aqueous NaHCO.sub.3. This solution is then added immediately to a C. g reverse phase column, eluted with water, and then the appropriate fractions are lyophilized to give 0.25 g of a light yellow solid. This material is again purified by reverse phase HPLC to give the product (0.210 g, 55%) as a light yellow solid.

IR (KBr) >>"* 3400 (br, -OH), 1755 (β-lactam CO), 1590 (-CO ₂ ") cm ^{-1 1} H-NMR (D ₂ O):: 8.60-7 , 76 (m, 4H, arom.), 4.76 (t, J =

5.8 Hz, 2H, N-CH ₂ ), 4.13 (d of q, J = J '= 6.3 Hz, 1H, H-1 ¹ ), 3.95 (d of t, J = 9 , 0 Hz, J '= 2.8 Hz, 1H, H-5), 3.45-2.75 (m, 5H), 2.59 (s, 3H, S-Me), 1.20 (i.e. , J = 6.4 Hz, 3H, CHMe) UV (H ₂ O): A _max : 296 U 8509), 273 U 13005), 231 (£ 11576) nm ^ _1/2 (pH 7.4, 36, 8 ° C) = 20 h.

Example 10

Preparation of 3- [2- (1- (2,6-dimethylpyridinium)) ethylthio] -6cc [1 - (R) -hydroxyethyl] -7-oxo-1-azabicyclo [3.2.0] -hept -2 -en-2-carboxylate

COO

A. 1 - (2-Mercaptoethyl) -2,6-dimethylpyridinium methane sulfonate

+ MsOH

100 ⁰ C, 42 h

MsO

A mixture of 2,6-dimethylpyridine (19.2 mL, 0.165 mol) and methanesulfonic acid (3.27 mL, 0.05 μ mol) is stirred for 15 min, treated with ethylene sulfide (4.17 mL, 0.070 mol) and stirred 42 h under nitrogen atmosphere at 100 ° C. After cooling to 25 ^° C., the reaction mixture is diluted with ether (45 ml) and water (30 ml), separating the two

20 25 30

Layers and extract the organic layer with water (2x5 ml). The aqueous layers are combined, filtered through a pad of celite, washed with ether (2 × 15 ml), traces of organic solvents are removed in vacuo and poured onto a U-Bondapack G-18 column (3.0 × 12 cm). , Elute with a 3% acetonitrile-97: 6 water mixture and lyophilize the appropriate fractions to give 2.5 g of the impure title compound as a syrup. This syrup is again purified by HPLC (/ U Bondapak C-18) to give 0.90 g (7%) of the title compound

IR (movie) \? _fflax : 2520 (SH), 1640 and 1625 (pyridinium),

1585, 1490, 1200 cm ^"1 (sulfonate) ¹ H-NI-IR (DMS0-d ₆ + D _£ 0) Sx 2.36 (3H, s, CH ₃ SO _3" "), 4.62

(2H, m, CH ₂ N ⁺ ), 7.74 (2H, m, · Hm of pyridinium),

8.24 (1H, m, Hp of pyridinium)

UV (H _o 0) A ", _QV : 272 (£ 4080) m / u

d. Max /

B. p-Nitrobenzyl-3- [2- (1- (2,6-dimethylpyridinium)) ethylthio] -6a [i - (R) -hydroxyethyl] -7-oxo-1-azabicyclo- Γ5.2.0] hept-2-ene-2-carboxylate-diphenylphosphate

1) NEt (iPr) ₂

2) ClP (OPh) ₂

OH

4) NEt (IPr) ₂

COOPNB

35

COOPNB

(PhO) ₂ PO "

To a cold ⁽⁰ ° C) solution of p-nitrobenzyl 6cc- [i- (R) -hydroxyethyl] -3, 7-dioxo-1-azabicyclo [3.2.0] heptane-2-carboxylate (0.658 g, 1 , 89 mmol) in 6 ml of acetonitrile (stored under nitrogen atmosphere) are added diisopropylethylamine (0.394 ml, 2.26 mmol) and diphenylchlorophosphate (0.468 ml, 2.26 mmol), the reaction mixture is stirred for 30 min and treated with a solution of 1 - (2-Mercaptoethyl) -2,6-dimethylpyridinium methanesulfonate (0.720 g, 2.73 mmol) in 3 mL acetonitrile and then with diisopropylethylamine (0.394 mL, 2.26 mmol). The reaction mixture is stirred at ⁰ ° C. for 2 h, Dilute with cold ( ⁰ C) water (27 mL) and pour onto a / U Bondapak C-18 column (2.5 x 9 _f 0 cm). Elute with an acetonitrile-water mixture and lyophilize the appropriate fractions to give 0.92 g (65%) of the title compound.

IR (KBr) v>: 3700-3000 (OH), 1765 (C = O of β-lactam), 1690 (C = O of the PNB ester), 1620 (pyridinium), 1590 (phenyl), 1517 (NO ₂ ), 1330 "(NO ₂ X 880 cm ^-1 NO ₂ ) ¹ H-NMR (DMSO-d ₆ ) S : 1.15 (3H, d, J = 6.2 Hz, CH ₃ CHOH), 2.7 -3.7 (11H, CH ₂ S, 2-CH ₃ on pyridinium, H-4, H-6), 3.7-4.4 (2H, CH ₃ CHOH, H-5), 4.7 ( 2H, m, CH ₂ N ^+), 5.14 (1H, d, J = 4.5 Hz, OH), 5.37 (center of ABq, J = 13.2 Hz, _CH2 of PNB), 6 , 7-7.5 (1OH, m, phenyl), 7.5-8.7 (7H, pyridinium, H of PNB) UV (H ₂ O) * _max : 274 (^ 14150), 319 (ε 9445) m / U

C. 3- [2- (1- (2,6-Dimethylpyridinium)) ethylthio] -6cc [1 (R) -hydroxyethyl] -7-oxo-1-azabicyclo [3.2.0] hept-2 - en-2-carboxylate 35

1 0% Pd / c,

THF, ether, buffer

COOPNB

(PhO) ₂ PO '

To a solution of p-nitrobenzyl-3- [2- (i- (2,6-dimethylpyridinium)) ethylthio] -6oc- [1- (R) -hydroxyethyl] -7-oxo-1-azabicyclo [3.2.0 ] Hept-2-ene-2-carboxylate-diphenylphosphate (0.80 g, 1.07 mmol) in humid tetrahydrofuran (42 mL) is added ether (42 mL), potassium phosphate monobasic sodium hydroxide buffer (0.15 M, pH 7.22, 21 mL) and 10% palladium on carbon (0.80 g), the resulting mixture is hydrogenated for 1 h at 23 ⁰ C and a pressure of 2.76 bar (40 psi) and filtered through a Celite -Pillow. Separate the two layers and extract the organic layer with the buffer (3x8 ml). The aqueous phases are combined, washed with 50 ml of ether, traces of organic solvents are removed in vacuo and added to a / U Bondapak C-18 column (3.0 × 10.2 cm). The column is eluted with a 5% acetone tris-95% water mixture and the appropriate fractions are lyophilized to give 0.246 g (63%) of the title compound as a yellowish powder. IR (KBr) C?

Max

3700-2800 (OH), 1750 (C = O of β-lactam),

-1

1620 (pyridinium), 1585 cm (carboxylate)

¹ H NMR (D ₂ O) i : 1.23 (3H, d, J = 6.4Hz, CH ₃ CHOH), 2.5-3.5 (11H, H-4, H-6, CH _£ S, 2CH, on pyridinium),

1020

3.8-4.4 (2H, CH ₃ CHOH, H-5), 4.5-4.9 (CH ₂ N ⁺ , HOD), 7.64 and 7.74 (2H, Α part of A ₂ B-Systems, Hm of pyridinium), 8.07, 8.16, 8.18 and 8.27 (1H, B part of the A ₂ B system, Hp of pyridinium) UV (H ₂ O) A _max : 277 (£ .9733), 300 ( t 8271) m _{/ U} [<x] Jp = + 50.7 ° (C 0.48, H ₂ O) Analysis: for C ₁ gH ^^ O ^ S. 1.5 H ₂ O

Calculated: C 55.51% H 6.47% N 7.19% Found: 55.14 6.23 6.46.

B ice ρ ie 1

11

Preparation of (5R, 6S) -3- [2- (2-methylthio-3-methylimidazolio) ethylthio] -6- [i- (R) -hydroxyethyl] -7-oxo-1-aza- mcyclo Γ 3.2. 0 lhept-2-ene-2-carboxylate

N-Me

A. 2-Methylthio-3-methyl-; 1- (2-mercaptoethyl) imidazolium trifluoromethanesulfonate

_v> HSCH ₂ CH ₂ N ..N-Me

SMe

CF ₃ SO ₃ V

Trifluoromethanesulfonic acid (1.38 ml, 0.015 mol) is added dropwise to 2-methylthio-1-methylimidazole (4.0 g, 0.03 mol) (prepared as described by A. Wohl and W. Marckwald in Chem. Ber.22 , 1353 (1889) described) at ^{0 0} C under

N ₂ , then ethylene sulfide (0.9 ml, 0.015 mol) is added, the mixture is warmed to 55 ° C under Np for 24 h, the reaction mixture is triturated with ether (3x) and the residue taken up in acetone. Then filtered and evaporated. This gives the product (4.2 g, 82%) as a semi-crystalline solid, which is used without further purification. IR (Film) v>: 2550 (black, Sch.) Cm ^{-1 1} H-NMR (d ₆ -acetone) S: 7.97 (s, 2H), 4.66 (t, J = 7 Hz, 2H , Methylene), 4.17 (s, 3H, N-Me), 3.20 (d of t, J = 7Hz, J '= 9Hz, 2H, methylene), 2.72 (s, (3H, S-Me), 2.20 (t, J = 9 Hz, 1H, -SH).

For example, p-nitrobenzyl- (5R, 6S) -3- [2- (2-methylthio-3-methylimidazolio) -ethylthio] -6- [i- (R) -hydroxyethyl] -7-oxo-iazabicyclo [3.2. 0] he "pt-2-ene-2-carboxylate-drphenyl phosphate

SMe

O ₂ PNB

To a solution of p-nitrobenzyl- (5R, 6S) -6- [i- (R) -hydroxyethyl] -3,7-dioxo-1-azabicyclo [3.2.0] heptane-2-carboxylate (1.40 g , 4.0 mmol) in 50 ml of dry acetonitrile is added under N ₂ at ^{0 0} C dropwise diisopropylethylamine (0.76 ml, 4.4 mmol) and then diphenylchlorophosphate (0.91 ml, 4.1 mmol), stirred Reaction mixture for 1 h at room temperature, gives diisopropylethylamine (0.76 ml, 4.4 mmol) and then a solution of 2-methylthio-3-methyl-1- (2-mercaptoethyl) imidazolium trifluoromethanesulfonate (2.0 g, 5.9 mmol) in 5 ml of acetonitrile dropwise, the reaction mixture keeps

Stir for 1.5 h at room temperature and then concentrated in vacuo to obtain a gummy product. This gummy product is taken up in water and placed on a C ^ o reverse phase column. It is eluted with water, then 20% acetonitrile-water and finally 30% acetonitrile-water. Then the appropriate fractions are lyophilized and the product (0.90 g, 30%) as a pale yellow solid.

IR (KBr) v ^? - 338O (br, OH), 1770 (β-lactam CO) cm " ¹ - max

¹ H-NMR (dg-acetone) S : 8.35 (brs, 1H, 8.24, 7.78 (ABq, J = 8.8 Hz, 4H, aromat.), 7.89 (brs, 1H) , 7.25-6.91 (m, 10H, diphenyl phosphate), 5.50, 5.25 '(ABq, J = 12Hz, 2H, benzyl.), 4.75-4.27 (m, 3H). , 4.03 (s, 3H, N-Me), 4.15-2.75 (m, 8H), 2.59 (s, 3H, S-Me), 1.22 (d, J = 6, 2 Hz, 3H, -CHMe).

C. (5R, 6S) -3- [2- (2-methylthio-3-methylimidazolio) ethylthio] -6- [i- (R) -hydroxyethyl] -7-oxo-1-azabicyclo-

Γ3.2.0] hept-2-ene-2-carboxylate

SMe.

CO ₂ PNB

(0 O) ₂ PO ⁰

To a solution of p-nitrobenzyl- (5R, 6S) -3- [2- (2-methylthio-3-methylimidazolio) -ethylthio] -6- [1- (R) -hydroxyethyl] -7-oxo-1 azabicyclo [3.2.0] hept-2-ene-2-carboxylate diphenyl phosphate (1.20 g, 1.56 mmol) in a mixture of 70 mL of THF, 70 mL of ether, and 31 mL of phosphate buffer (0.05 M , pH 7.4) is added 1.2 g of 10% palladium on carbon, the mixture (Parr) is hydrogenated for 55 min at a pressure of 2.41 bar (35 psi), the reaction mixture is filtered through Celite and washed the filter cake with water and ether. The aqueous phase is separated, cooled to ^{0 0} C and the pH with saturated, aqueous NaHCO, to 7.0. After removal of residual organic solvents in vacuo, the aqueous solution is added to a C ^ g reversed-phase acid. It is eluted with water and then with 8% acetonitrile-water and then the appropriate fractions are lyophilized to give 0.25 g of a solid. This material is again purified by reverse phase HPLC to give the product (0.114 g, 19%) as an off-white solid.

IR (KBr) 0 _m -. 3420 (OH), 1750 (β-lactam CO), .1590 (-CO ₂ ") cm ^-1

¹ H-NMR (B ₂ O) S: 7.58 (s, 2H), 4.52 (t, J = 6Hz, 2H), 4.28-3.82 (m, 2H), 3.90 (s, 3H, N-Me), 3.40-2.87 (m, 5H), 2.40 (s, 3H, S-Me), 1.20 (d, J = 6.4Hz, 3H , -CHMe) UV (H ₂ O) X _max : 297 (£ 7572), 262 (£ 6,259), 222 (£ 7,955) mn.

example

Preparation of (5R, 6S) -3- [2- (3-aminopyridinio) ethylthio] -6- [1 - (R) -hydroxyethyl] -7-oxo-1-azabicyclo [3.2.0] .hept -2 -en-2-carboxylated t

A. 3-Amino-1- (2-mercaptoethyl) -pyridinium chloride

.HCl

NH

HSCH ₂ CH ₂

Cl

3-Aminopyridine (1.50 g, 0.016 mol) is taken up in 15 ml of 1N methanolic HCl and the resulting solution is evaporated to give the hydrochloride as an oil, giving 3-aminopyridine (1.32 g, 0.015 mol) and ethylene sulfide (0.39 ml, 0.015 mol) of this oil and heated the resulting mixture (oil bath) for 2 h under N ₂ at 60 to 65 ° C. Another equivalent of ethylene sulfide (0.89 ml, 0.015 mol) is added and the mixture is heated for a further 65 h at 55 to 65 ^° C. The reaction mixture is washed with CH ₂ Cl ₂ and then taken up in 25 ml of water. The aqueous solution is added to a co-reversed phase column, which is eluted with water. After evaporation of the relevant fractions, the product (1.26 g, 44%) is obtained as a colorless, viscous oil.

IR (film) O _m ι 3180 (NH ₀ ) cm ^-1 , max <L

¹ H-NMR (dg-DMSO) C 8.19 to 7.59 (m, 4H, aromat.), 4.59 (t, J = 6.2 Hz, 2H, N-CH _2), 3.5 (brs, 2H, -NH ₂ ), 3.20-2.77 (m, 3H).

B. p-Nitrobenzyl- (5R, 6S) -3- [2- (3-aminopyridinio) -ethylthio] -6- [1- (R) -hydroxyethyl] -7-oxo-1-azabicyclo [3.2.0] hept-2-ene-2-carboxylate dipheny! phosphate

CO ₂ PNB

CO ₂ PNB

To a solution of p-nitrobenzyl- (5R, 6S) -6- [1- (R) -hydroxyethyl] -3,7-dioxo-1-azabicyclo [3.2.0] heptane-2-carboxylate (0.696 g, 2 , 0 mmol) in 10 ml of dry acetonitrile are added under N ₂ at ^{0 0} C dropwise diisopropylethylamine (0.382 ml, 2.2 mmol) and then diphenylchlorophosphate (0.457 ml, 2.2 mmol), stirred for 30 min at 0 ⁰ C, Add a solution of 3-amino-1- (2-mercaptoethyl) -pyridinium chloride (0.475 g, 2.5 mmol) in 1 mL of dry DMF and then add more diisopropylethylamine (0.435 mL, 2.5 mmol), keeping the reaction mixture 1 , 5h at ^{0 0} C and then concentrated in vacuo. The resulting gummy product is taken up in acetonitrile-water (1: 1) and added to a co-reversed phase column. It is eluted with water, then with 20% acetone tril water and then the relevant fractions are lyophilized to give the product (0.730 g, 50%) as a beige solid.

IR (KBr)? : 3330 (br, OH), 3180 (br, NH ₂ ), 1770 (β-Lac

tam CO), 1690 (-CO ₂ PNB) cm " ¹

¹ H-NMR (dg-DMSO) 6 : 8.29-7.63 (m, 8H, aromat.), 7.2-6.7 (m, 1 OH, diphenylphosphate), 5.47, 5, 18 (ABq, J = 14Hz, 2H, benzyl.), 4.73-4.45 (m, 3H), 4.2-3.8 (m, 1H), 3.6-2.6 (m , 8H), 1.15 (d, J = 6.2 Hz, 3H), CHMe)

10

C. (5R, 6S) -3- [2- (3-aminopyridinio) ethylthio] -6- [i (R) -hydroxyethyl] -7-oxo-1-azabicyclo [3.2.0] hept-2 en-2- carboxylate ·

15 2-0 25 30

OH

35

To a mixture of p-nitrobenzyl- (5R, 6S) -3- [2- (3-aminopyridinio) -ethylthio] -6- [1- (R) -hydroxyethyl] -7-oxo-1-azabicyclo [3 2.0] hept-2-ene-2-carboxylate diphenyl phosphate (0.730 g, 1.0 mmol) and 10% palladium on carbon (o, 7 g) in 25 ml of a phosphate buffer (0.05 M, pH 7.4) are added 8 ml of THF and 20 ml of ether, the mixture is then hydrogenated for 1 h at a pressure of 2.76 bar (40 psi) (Parr), the mixture obtained filtered through a pad of Celite and washed with the filter cake Water and ether. The

aqueous phase is separated off, washed twice with ether and then removed residual volatiles in vacuo. The aqueous solution is immediately added to a C ₁₈ reverse phase column, which is eluted with water. Lyophilization of the relevant fractions gives 0.45 g of an off-white solid. This material is again purified by reverse phase HPLC to give the desired product (0.123 g, 35%) as an off-white solid

IR (KBr) v? _max : 3340 (br), 1750 (br, β-lactam CO), 1580 (br, -CO ₂ ") cm

¹ H-NMR (D ₂ O) Si 8.07-7.59 (m, 4H, aromat.), 4.61 (t, J = 5.8Hz, 2H, N-CH ₂ ), 4.14 (d of q, J = J '= 6.3 Hz, 1H, H-1 ¹ ), 3.97 (d of t, J = 9.2 Hz, J' = 2.6 Hz, 1H, H- 5), 3.38 (t, J = 5.8 Hz, 2H, S-CH ₂ ), 3.24 (d of d, J = 6.0 Hz, J = 2.6 Hz, 1H, H -6), 3.17-2.57 (m, 2H, H-4), 1.21 (d, J = 6.3Hz, 3H, CHMe) UV (H ₂ O) * _max : 299 (£ 7949), 256 (£ 8822) T. nm _{/ o} (pH 7.4, 36.8 ⁰ C) = 18.5 h.

Example 13 Preparation of

(5R, 6S) -3- [i- (S) -methyl-2- (1-pyridinium) ethylthio] -6- [1 - (R) -hydroxyethyl] -7-oxo-1-azabicyclo [3.2. 0] hept-2- ene-2-carboxylate

and

(5R, 6S) -3- [i- (R) -methyl-2- (1-pyridiniujn) ethylthio] -6- [1- (R) -hydroxyethyl] -7-oxo-1-azabicyclo [3.2. 0] hept-2- ene-2-carboxylate

1015

coo

A. dl-1 - (2-mercapto-2-methylethyl) -pyridini \ Jin-niethan-

sulfonate

dl-1 - (2-mercapto-i-methylethyl) pyridinium methanesulfonate

SH

MsOH

Methanesulfonic acid (1.95 ml, 0.030 mol) is added slowly to cold pyridine (7.83 ml, 0.097 mol), the resulting mixture is stirred for 15 min at 40 ⁰ C, treated with dl ^ Propylensulfid (2.59 ml, 0.033 And stirred for 90 h under a nitrogen atmosphere at 60 ⁰ C. The pyridine is removed in vacuo, the residue is mixed with Y / asser and purified by chromatography (HPLC, Preparative Bondapak C-18). The appropriate fractions are combined and lyophilized to give dl-1- (2-mercapto-2-methylethyl) -pyridinium methanesulfonate (1.14 g, 15%) as a colorless syrup.

35

IR (film) ^ _max : 2520 (SH), 1640 (pyridinium), 1180 (st,

CH ₃ SO ₃ "), 1040 (CH ₃ SO ₃ ") cm ^-1

¹ H-NMR (DMSO-d ₆ ) <5: 1.35 (d, J = 6.8 Hz, 3H, CH ₃ CHS), 2.30 (s, 3H, CH ₃ SO ₃ "), 2, 90 (d, J = 8.5 Hz, 1H, SH), 3.2-3.7 (m, CHSH), 4.52 (dd, J = 12.9 Hz, J = 8.4 Hz, CHCH ₂ N), 4.87 (dd, Jg _81n = ^12, 9 Hz, J = - 6.0 Hz, CHCH ₂ N ^+), 8.0-8.4 (m, 2H, Hm of pyridinium), 8 , 5-8.8 (m, 1H, Hp of pyridinium), 9.04 (dd, J = 1.4Hz, J = 6.7Hz, 2H, Ho of pyridinium) UV (H ₂ O) ^ _max : 208 (£ 5267), 259 (£ 3338) Analysis: for

Calculated: C 37.88% H 6.71% N 4.91% S 22.47% Found: 37.49 6.85 4.86 22.09

Further, dl-1- (2-mercapto-1-methylethyl) -pyridinium methanesulfonate (0.82 g, 11%) is obtained as a colorless syrup.

IR (film) w> ": 2500 (SH), 1628 (pyridinium), 1180 (SuI)

fonate), 1035 (sulfonate) cm ^-1 H-NMR (DMSO-d ₆ ) S: 1.69 (d, J = 6.8 Hz, 3H, CH ₃ CHN ⁺ ), 2.31 (s, 3H, CH ₃ SO ₃ "), 3.0-3.3 (m, 2H, CH ₂ S),

4.2-5.2 (m, 1H, CHN ⁺ ), 8.0-8.4 (m, 2H, Hm of pyridinium), 8.5-8.8 (m, 1H, Hp of pyridinium),

9.0-9.2 (m, 2H, Ho of pyridinium) UV (H ₂ O) X _max : 209 (L 4987), 258 (6 3838) Analysis: for C ₉ H ₁₅ NO ₃ S ₂ .1, 5H _£ 0

Calculated: C 39.11% H 6.56% N 5.07% Found: 39.13 5.92 5.20

B. (5R, 6S) -p-nitrobenzyl-3- [1- (R, S) -methyl-2- (1-pyridinium) -thyrithio] -6- [i- (R) -hydroxyethyl] - 7-oxo-1-aza- bicyclo Γ 3.2.0] hept-2-ene-2-carboxylate-diphenylphosphate 35

OH

1) NEt (IPr) ₂

'Ί-Γ ^ \ _ ² ^ ^clp (° ^Ph )

4) NEt (iPr).

10 15 20 25 30

COOPNB

To a cold (0 ° C) dissolution of (5R, 6S) -p-nitrobenzyl-6- [1- (R) -hydroxyethyl] -3,7-dioxo-1-azabicyclo [3.2.0] heptane-2 Carboxylate (0.523 g, 1.5 mmol) in 6 mL of acetonitrile (stored under nitrogen atmosphere) are added diisopropylethylamine (0.314 mL, 1.8 mmol) followed by diphenylchlorophosphate (0.373 mL, 1.8 mmol), the reaction mixture is stirred for 30 min and treated with a solution of dl-1- (2-mercapto-2-methylethyl) -pyridinium methanesulfonate (0.539 g, 2.16 mmol) in 2 mL of acetonitrile and diisopropylethylamine (0.314 mL, 1.8 mmol). The reaction mixture is stirred for 1 h at ^{0 0} C, diluted with cold (O ⁰ C) water (24 ml) and chromatographed on a prep.Bondapak C-18 column (2.5 x 8.5 cm), taking 23 % to 50% acetonitrile in water used as eluent. After lyophilization, 1.07 g (97%) of the title compound are obtained as a yellowish powder.

35

IR (KBr) \? _m ": 3700-3100 (OH), 1770 (C = O of β-lactam), 1695 (C = O of the PNB ester), 1630 (pyridinium), 1590 (phenyl), 1518 (NO ₂ ), 1348 ( NO ₂ ), 885 (NO ₂ )

cm

-1

¹ H-NMR (DMSO-dg) <£: 1.14 (d, J = 6.1 Hz, 3H, _CH3 CHO), 1.33 (d, J = 6.3 Hz, 3H, CH ₃ CHS ), 4.6-5.0 (m, CH ₂ N ⁺ ), 5.14 (d, J = 5.2 Hz, 1H, OH), 5.37 (center of ABq, J = 12.4 Hz , 2H, CH ₂ of PNB), 6.6-7.5 (m, 1OH, phenyl of the phosphate), 7.69 (d, J = 8.7 Hz, 2H, Ho of PNB), 8.0 8.4 (m, 4H, Hm of PNB, Hm of pyridinium), 8.4-8.8 (m, 1H, Hp of pyridinium), 9.08 (d, J = 5.6 H7, 2H, Ho of pyridinium) UV (H ₂ O) A _max : 263 (£ 13325), 308 (έ 8915) Analysis: for C ₃ H-ZgN ₃ O ₁₀

Calculated: C 57.52% H 5.1090 found: 57.76 4.96

.H ₂ O

N 5.59% S 4.27% 5.36 4.35.

C. (5R, 6S) -3- [i- (R and S) -methyl-2- (1-pyridinium) ethylthio] -6- [1 - (R) -hydroxyethyl] -7-oxo-1 azabicyclo [3.2.0] hept-2-ene-2-carboxylate

OH

COOPNB

10% Pd / C, H ₂

THF, ether, 0 buffer

(PhO) ₂ PO "

COO

To a solution of (5R, 6S) -p-nitrobenzyl-3- [i- (R, S) -methyl-2- (1-pyridinium) ethylthio] -6- [1- (R) -hydroxyethyl] -7-oxo-1-azabicyclo [3.2.0] hept-2-ene-2-carboxylate diphenyl phosphate (0.60 g, 0.82 mmol) in humid tetrahydrofuran (33 ml) is added ether (33 ml), potassium phosphate monobasic sodium hydroxide Buffer (17 ml, 0.15 M, pH 7.22) and 10 g of palladium on carbon (0.60 g) and hydrogenates the resulting mixture for 1 h at 23 ⁰ C and a pressure of 2.76 bar ( The two layers are separated and the organic layer extracted with water (3x7 ml), the aqueous layers combined, filtered through a pad of celite, washed with ether (3 × 20 ml) and chromatographed on a prep.Bondapak C-18 column (2.5 x 9 »5 cm) using water as the eluting solvent to give 0.18 g (63% of a mixture of diastereoisomers) The two diastereoisomers are separated by HPLC (prep.Bondapak C, J. Med -18) using water as the eluent t 0.068 g (23%) of the lower retention time isomer, designated Compound "B".

IR (KBr) V _η 1770 (C = O of β-lactam), 1633 (pyridinium), 1593 (carboxylate) cm "

¹ H-NMR (D ₂ O) S: 1.20 (d, J = 6.3 Hz, 3H, CH ₃ CHO), 1.42 (d, J = 6.9 Hz, 3H, CH ₃ CHS) , 2.3-3.2 (m, 3H, H-4, H-6), 3.5-3.9 (m, 1H, SCH), 3.9-4.2 (m, 2H, H -5, CH ₃ CHO), 4.3-5.1 (m, CH ₂ N "), 7.8-8.2 (m, 2H, Hm of pyridinium), 8.4-8.7 (m , 1H, Hp of pyridinium), 8.7-9.0 (m, 2H, Ho of pyridinium) UV (H ₂ O) \ _max : 260 (£ 6727), 300 (£ 8245) ίαψ = -39.3 ° (c, H ₂ O)

^ 1 / ₌ 12.6 (measured at a concentration of 10 ^"4 M in phosphate buffer pH 7.4 at 36.8 ⁰ C) h.

30

The isomer having a higher retention time is further obtained as 0.081 g (28%), designated Compound "A".

IR (KBr) ^ _max : 1755 (C = O of β-lactam), 1630 (pyridinic)

um), 1590 (carboxylate) cm

H-NMR

(D ₂ O) S x 1.18 (d, J = 6.3 Hz, 3H, CH ₃ CHO), 1.40 (d, J = 7.0 Hz, 3H, CH ₃ CHS), 2.84 ( d, J = 9.3Hz, 2H, H-4), 3.26 (dd, J = 2.7Hz, J = 5.9Hz, 1H, H-6), 3.4 - ^, 2 (m, 3H, SCH, CH ₃ CHO, H-5), 4.2 to 5.1 (m, CH ₂ N ^+), 7.7-8.1 (m, 2H, Hm of pyridinium), 8 , 3-8.65 (m, 1H, Hp of pyridinium), 8.65-8.9 (m, 2H, Ho of pyridinium) UV (H ₂ O) A _max : 259 (6 5694), 296 (ε 6936) [α] 23 ₌ + 96.9 ° (c 0.56, H ₂ O)

_T1 / ₂ = 15.6 h (measured at a concentration of 10 ~ ⁴ M in phosphate buffer pH 7.4 at 36.8 ⁰ C)

Example 14 Preparation of

(5R, 6S) -3- [2 - [(S) - (I -pyridinium)] -1- (S) -cyclohexylthio] -6- [1 - (R) -hydroxyethyl] -7-oxo-1 azabicyclo [3.2.0] hept-2- ene-2-carboxylate

35

and

(5R, 6S) -3- [2 - [(R) - (1-pyridinium)] - 1- (R) -cyclohexylthlo].

6- [i- (R) -hydroxyethyl] -7-oxo-1-azabicyclo [3.2.0] hept-2-e n-2-carboxylate

OH

A_

10

COO "

A. dl-1- (2-mercapto-1-cyclohexyl) pyridinium methanesulfonate

S + Ni 1> + MsOH> V ^ V. _r "\ MsO

Methanesulfonic acid (0.65 mL, 0.01 mol) is added dropwise to pyridine (2.42 mL, 0.03 mol) with cooling, the mixture is stirred for 10 min under a nitrogen atmosphere, treated with dl-cyclohexene sulfide [1.377 g (85 % purity), 0.0102 mol) and stirred .25 h at 72 ° C. Excess pyridine is removed in vacuo. Traces are distilled together with water. The residue is mixed with water and chromatographed on a prep.Bondapak C-18 column (5 × 13 cm) with 0-290 acetonitrile in water as eluent. After lyophilization, 1.57 g (53%) of a colorless syrup are obtained. IR (FiIm) O: 2500 (SH), 1625 (pyridinium), 1190 (SO-)

35

¹ H-IMR (DMSO-d ₆ ) S : 1.2-2.5 (m, 8H, cyclohexyl H), 2.32 (s, 3H-, CH ₃ SO ₃ "), 2.82 (d, J = -9.8 Hz, SH), 3.0-3.5 (m, 1H, CHSH), 4.2-4.9 (m, 1H, CHN ⁺ ),

8.0-8.3 (m, 2H, Hm of pyridinium), 8.4-8.8 (m, 1H, Hp of pyridinium), 8.9-9.3 (m, 2H, Ho of pyridinium) UV (H ₂ O) A _max : 214 (£ 5365), 258 (13500) Analysis: for C ₁₂ H ₁

Calculated: C 46.88% Η 6.88% N 4.56% found: 46.61 6.46 4.65

B. (5R, 6S) -p-nitrobenzyl-3- [2 - [(R or S) - (1-pyridinium)] • 1- (R or S) -cyclohexylthio] -6- [i- (R) -hydroxyethyl] -7-0x0-1-azabicyclo [3.2.0] hept-2-ene-2-carboxylate-di- phenyl phosphate

20

) NEt (iPr) ₂

0 2) ClP (OPh) ₂

COOPNB

4) NEt (iPr)

35

(PhO) ₂ PO '

To a cold ⁽⁰ ° C) solution of (5R, 6S) -p-nitrobenzyl-6- [1- (R) -hydroxyethyl] -3,7-dioxo-1-azabicyclo [3.2.0] heptane-2 carboxylate (1.37 g, 3.93 mmol) in 15 mL of acetonitrile (stored under a nitrogen atmosphere) are added diisopropylethylamine (0.822 mL, 4.7 mmol) and diphenylchlorophosphate (0.979 mL, 4.7 mmol). The resulting solution is stirred for 30 minutes, treated with a solution of dl-1 - (2-mercapto-1-cyclohexyl) pyridinium methanesulfonate (1.64 g, 5.66 mmol) in 4.7 ml of acetonitrile and then with diisopropylethylamine (0.822 ml, 4.7 mmol). The reaction mixture is stirred for 1 h at ⁰ ° C, diluted with cold ⁽⁰ ° C) water (75 ml) and chromatographyert using a präp.Bondapak C-18 column eluting using 25-50% acetonitrile in water as eluent. After lyophilization of the appropriate fractions, 1.9 g (53%) of the title compound are obtained.

IR (KBr) v>: 3700-3000 (OH), 1770 (C = O of β-lactam), 1700 (C = O of the PNB ester), 1628 (pyridinium), 1590 (phenyl), 1515 (NO ₂ ), 1345 (NO ₂ ), 880 (NO ₂ ) cm ^{-1 1} H-NMR (D ₂ O) S : 1.13 (d, J = 6.1 H 7, 3H, CH ₃ CHO), 1.2 -2.5 (m, 8H, cyclohexyl H), 2.7-3.5 (m, 4H, H-4, CHS), 3.5-4.4 (m, 2H, _CH3 CHO, H 5), 4.4-5.0 (m, 1H,

CHN ⁺ ), 5.30 (center of ABq, J = 12.8 Hz, CH ₂ of PNB), 6.7-7.4 (m, 1OH, phenyl), 7.65 (d, J = 8, 6Hz, 2H, Ho of PNB), 7.9-8.4 (m, 4H, Hm of PNB, Hm of pyridinium), 8.4-8.8 (m, 1H, Hp of pyridinium), 9, 0-9.4 (m, 2H, Ho of pyridinium) UV (H ₂ O) ^ _max : 263 (£ 9038), 309 (£ 6394) Analysis: for C ₃₀

Calculated: C 59.16% H 5.35% N 5.31% Found: 58.95 5.15 5.57

C. (5R, 6S) -3- [2 - [(R or S) - (i-pyridinium)] - 1- (R or S) -cyclohexylthio] -6- [i- (R) -hydroxyethyl] - 7-oxo-1-azabicyclo [5 »2.0] hept-2-ene-2-carboxylate

OH

• N

COOPNB ^. ¹¹

10% Pd / C, H _2, THF, ether, PuffSr

0 (PhO) ₂ PO "

20 · -

To a solution of (5R, 6S) -p-nitrobenzyl-3- [2 - [(R or S) - (I -pyridinium)] - 1- (R or S) -cyclohexylthio] -6- [i- ( R) -hydroxyethyl] -7-oxo-1-azabicyclo [3.2-0] hept-2-ene-2-carboxylate-diphenylphosphate (1.85 g, 2.34 mmol) in humid tetrahydrofuran (96 mL) Ether (96 mL), potassium phosphate monobasic sodium hydroxide buffer (0.15 M, pH 7.22, 50 mL) and 10% palladium on carbon (1.9 g) and the resulting mixture hydrogenated for 1.25 h at 23 ° C under a pressure of 2.76 bar (40 psi). The organic layer is separated, washed with water (3 × 20 ml), the aqueous solutions are filtered through a celite pad, washed with ether (2 × 60 ml) and traces of organic solvents are removed by pumping and chromatography on a prep Bondapak C-18 column (4.5 x 9 cm) with 0-5% acetonitrile in water as eluent. You get after

Lyophilization 0.705 g (76%) of a mixture of diastereoisomers. The separation of these diastereoisomers is carried out by HPLC (prep.Bondapak C-18) with 4% acetonitrile in water as the eluent. The lower retention time diastereoisomer is designated Compound "A" (0.29 g, 31%)

IR (KBr) 0 _mav s 1750 (C = O of β-lactam), 1620, (Sch, pyridinium), 1685 (carboxylate) cm

¹ H-NMR (D ₂ O) O: 1.21 (d, J = 6.3 Hz, 3H, CH ₃ CHO), 1.4-2.5 (m, 8H, cyclohexyl H), 2.5 -3.05 (m, 2H, H-4), 3.05-3.25 (m, 1H, H-6), 3.3-3.7 (m, 1H, CHS), 3.9- 4.3 (m, 2H, H-5, _CH3 CHO), 4.3-4.8 (m, CHN ^+), 7.8-8.2 (m, 2H, Hm of pyridinium), 8, 3-8.7 (m, 1H, Hp of pyridinium), 8.8-9.1 (m, 2H, Ho of pyridinium)

UV (H ₂ O) X _max : 260 (£ 7123), 300 (ε 8685) [ocJJP = + 6.2 ° (c 0.63, H ₂ O

T w ₂ = 16.6 h (measured at a concentration of 10 ~ ⁴ M in phosphate buffer pH 7.4 at 36.8 ⁰ C)

O ₄ S .2H ₂ O N 6 60% S 7 , 55% H 6 , 65% 6 59 7 , 43rd 6 , 47

Analysis: for

calculated: C 56.59%

found: 56.83 25

The higher retention time diastereoisomer is referred to as compound "B" (0.35 g, 38%). IR (KBr) v? _v : 1750 (C = O of β-lactam), 1622 (Sch, Pyridinium), 1588 (carboxylate) cm ^-1 H-NMR (D ₂ O) S: 1.19 (d, J = 6.4Hz, 3H, _CH3 CHO), 1.3-2.5 (m, 8H, cyclohexyl H), 2.5-3.1 (m, 2H, H-4), 3.1-3.3 (m, 1H, H-6), 3.3-3.8 (m, 2H, H-5, CHS), 4.1 (center of m, 1H, _CH3 CHO), 4.25 to 4.7 (m , 1H, CHN ⁺ ), 7.8-8.1 (m, 2H, Hm of pyridinium), 8.3-8.7 (m, 1H, Hp of pyridinium), 8.75-9.0 (m , 2H, Ho of pyridinium)

UV (H ₂ O) ^ _max : 259 (6 5992), 296 (£ 7646) [α] ²³ = + 65.3 ° (c 0.43, H _p 0

D 1/2

= 20.2 h (measured at a concentration of

₁₀

-4

p _{phosphate buffer pH}

_at

Example 15

A. (5R) -Allyl-3 - [(2-pyridinioethyl) -tMo] - (6S) - [(1R) -hydroxyethyl] -7-oxo-1-azabicyclo [3.2.0] hept-2-ene 2-carboxylate dipheny! phosphate

1) ClPO (Οφ)

ΌΡΟ (Οφ)

To a solution of (5R) -allyl-3,7-dioxo (6S) - [(iR) -hydroxyethyl] -1-azabicyclo [3.2.0] heptane (2R) -carboxylate (473 mg, 1.87 mmol) in 6 ml of CH ₃ CN is added at about -10 ° C under nitrogen atmosphere diisopropylethylamine (0.42 ml, 2.4 mmol) and then diphenylchlorophosphate (0.50 ml, 2.4 mmol). The mixture is stirred at ⁰ ° C. for 30 minutes, cooled to -15 ° C., and an oily suspension of N- (2-mercaptoethyl) -pyridinium chloride (527 mg, 3.00 mmol) in 1 ml of CH ₅ CN is added of 5 drops of DMF followed by diisopropylethylamine (0.42 ml, 2.4 mmol).

The mixture is stirred at -15 C for 30 min, diluted with 20 ml of water, this mixture is purified directly on a reverse phase silica gel column (C ^ ₈ PrepPAK, 12 g, Waters Associates) eluting with water (200 ml) 10% CTUCN / H ₂ O (100 ml), 20% CH ₃ CN / H ₂ O (100 ml), 30% CH ₃ CN / H ₂ 0 (100 ml) and then with 40% CH ₃ CN / H ₂ 0 (100 ml ). The appropriate fractions are collected, the organic solvent removed by a vacuum pump and lyophilized to give 786 mg (1.26 mmol, Yield 67.3%) of the title compound as a brownish powder. ¹ H-NMR (DMSO-d ₆ , CFT-20) S : 1.16 (3H, d, J = 6 Hz, 1'-CH ₃ ),

2.6-3.7 (m), 3.75-4.3 (2H, m, 5-H and 1'-H), 4.65 (2H, m, -CO ₂ CH ₂ -), 4 , 87 (2H, t, J = 6 Hz,

-CH ₂ -N ⁺ ), 5-6,2 (3H, m, olefinic protons), 6,6-7,4 (m, aromat. Protons), 8,15 (2H, t, J = 7 Hz, aromat.protons meta to nitrogen), 8.63 (1H, t, J = 7 Hz, aromat.Proton para to nitrogen), 9.07 ppm (2H, d, J = 7 Hz, aromatic, proton ortho to nitrogen )

IR (film) 0 : 3400 (OH), 1770 (β-lactone), 1690 (ester), 1625 (pyridinio).

For example, (5R) -3 - [(2-pyridinioethyl) thio] - (6S) - [(1R) -hydroxyethyl "1-7-OXQ-1- azabicyclor3" 2.0 "~ hept-2-ene-2 carboxylate

To a solution of (5R) -allyl-3 - [(2-pyridinioethyl) thio] - (6S) - [(iR) -hydroxyethyl] -7-oxo-1-azabicyclo [3.2.0] hept-2 -en-2-carboxylate diphenyl phosphate (156 mg, 0.25 mmol) in 2 ml CH CN ₁₅ was subsequently added, at about 22 ⁰ C, a solution of potassium 2-ethylhexanoate in EtOAc (0.5 M, 0, 6 ml;! 0.3 mmol), Tripheny phosphine (15 mg, _t 0.057 mmol) and tetrakistriphenylphosphine palladium (15 mg, 0.013 mmol) and the mixture is stirred 2 h at about 22 ⁰ C under a nitrogen atmosphere. After adding 7 ml of anhydrous Et ₂ O, the precipitate is filtered, washed with 7 ml of anhydrous Et ₂ O and dried in vacuo to give 101 mg of a brownish solid. This is purified by reverse phase column chromatography (C ₁₈ PrepPAK, 12 g, Waters Associates) eluting with water. Suitable fractions (Fr.7-12, 20 ml each) are collected and lyophilized to give 53 mg (0.16 mmol, yield 64%) of the title compound as a yellowish powder. This material is contaminated with potassium diphenyl phosphate and potassium 2-ethylhexanoate

¹ H-NMR (D ₂ O, CFT-20) <S: 0.80 (t, J = 6.4 Hz, Me of ethylhexanoate), 1.21 (3H, d, J = 6.3 Hz, 1 'Me), 2.93 (2H, dd, 3 _Λ _ ₅ = 9 Hz, J _gem = 4 Hz, 1-Hs),

3.28 (1H, dd, Jg _-1 = 6.2 Hz, Jg_ ₅ = 2.5 Hz, 6-H), 3.42 (2H, t, J = 6 Hz, -CH ₂ S), 3 , 98 (1H, td, J _5-1 = 9 Hz, J ₅ _ ₆ = 2.5 Hz, 5-H), 4.15 (1H, q, J = 6.2 Hz, 1'-H) , 4.80 (2H, t, J = 6.0 Hz, -CH ₂ N ⁺ ), 7-7.5 (m, phenylphenyls of diphenylphosphate), 8.03 (2H, m, Hm of pyridinium) , 8.56 (1H, m, Hp of pyridinium) and 8.81 ppm (2H, d, J = 6.5 Hz, Ho of pyridinium).

Example 16

Preparation of 3- [2- (N-methylthiomorpholinium) ethylthio] -6a [i '- (R) -hydroxyethyl] -7-oxo-1-azabicyclo [3-2.O] -hept-2 ene-2-carboxylate

A. N-methyl-N- (2-mercaptoethyl) thiomorpholinium methanesulfonate

MeN

1) MsOH

²⁾ Δ

MsO

To pre-cooled (ice bath) N-methylthiomorpholine (5.00 g, 42.7 mmol) (sJMLehn and J.Wagner, Tetrahedron, 26 , 4227 (1970)) is added methanesulfonic acid (1.47 ml,

20.5 mmol) and ethylene sulfide (1.30 ml, 21.4 mmol), the mixture is heated to 65 ° C for 24 h and diluted with 25 ml of water. The aqueous solution is washed with diethyl ether (3 x 25 ml), pumped under vacuum and poured onto a silica gel reverse phase column. The title compound is eluted with water. The appropriate fractions are combined and concentrated to give the thiol as an oil (4.80 g, 86% yield).

IR (FiIm) J _{m Q} ": 2550 cm" ¹ (bl, SH), max.

¹ H-NMR (DMSO-d ₆ ) S: 3.25-2.95 (6H-, m, CH ₂ N), 3.32 (3H, s, CH ₃ N ⁺ ), 3.20-2, 65 (7H, m, CH ₂ S, SH) and 2.32 ppm (3H, s, CH ₃ SO ₅ ).

B. p-Nitrobenzyl-3- [2- (N-methyl-thiomorpholinium-diphenyl-.phosphate) ethylthio] -6 <x- [i '- (R) -hydroxyethyl] -7- oxo-1-azabicyclo [ 3.2.0] hept-2-ene-2-carboxylate

CO ₂ PNB

D EtN (-0

2). ClPO (Οφ) 3)

CO PNB OP (O0).

* ·it z

A cold (ice-bath) solution of p-nitrobenzyl-6a [i '- (R) -hydroxyethyl] -3,7-dioxo-1-azabicyclo [3.2.0] heptane-2-carboxylate (557mg, 1.60 in 8 ml of CH, CN are treated dropwise with diisopropylethylamine (0.336 ml, 1.92 mmol) and diphenylchlorophosphate (0.400 ml, 1.92 mmol), stirred for 30 min, the reaction mixture is treated with N-methyl-N- (2 -mercaptoethyl) thiomorpholiniimethanesulfonate (893 mg, 2.29 mmol) in 4 mL of CH 2 CN and with diisopropylethylamine (0.336 mL, 1.92 mmol) and stirred for 30 min. The solution is diluted with 20 ml of water and poured onto a silica gel reverse phase column. The desired compound is eluted with a 50% acetonitrile-water mixture, the appropriate fractions combined, pumped under vacuum for 2 h and lyophilized to give the title compound (1.01 g,

-1

IR (Nuool) \ 7 _m _ "1760 (s, .beta.-lactam C = O), 1510 cm" st ⁿ (,

Max

-1

NO ₂ ) cm

¹ H NMR (DMSO-dg) S: 8.25 (2H, d, J = S, 8Hz, H-aromat.), 7.70 (2H, d, J = 8.8Hz, H-aromat ), 7.33-6.84 (1OH, m, H-arom.), 5.37 (2H, center of ABq, J = 14.2 Hz, CH ₂ ), 5.14 (IH, d, J = 4.5Hz, OH), 4.35-3.80 (2H, m, H-1 x and H-5), 3.75-3.45 (6H, m, CH ₂ N ⁺ ), 3.31 (3H, s, CH ₃ N ^+), 3.45 to 2.75 (9H, m, CH ₂ S, H-6 and H-4) and 1.15 ppm (3H, d, J = 6.2 Hz, CH,).

15

25 30

C. 3- [2- (N-Methyl-thiomorpholinium) ethylthio] -6a [i '(R) -hydroxyethyl] -7-oxo-1-azabicyclo [3.2.0] hept-2- ene-2 carboxylate

A solution of p-nitrobenzyl-3- [2- (N-methyl-thiomorpholinium-diphenylphosphate) ethylthio] -6a [i '- (R) -hydroxyethyl] -7-oxo-1-azabicyclo [3.2. 0] hept-2-ene-2-carboxylate (1.31 g, 1.76 mmol) in 0.1 M pH 7.4 phosphate buffer

(48.8 ml), tetrahydrofuran (20 ml) and diethyl ether (20 ml) are hydrogenated for 1 h at 40 psi over 10% Pd / C (1.5 g) in a Parr shaker. The reaction mixture is diluted with 40 ml of diethyl ether and the phases are separated. The organic phase is washed with water (2 × 5 ml), the aqueous phases are combined, filtered through a # 52 hardened filter paper, washed with diethyl ether (2 × 20 ml) and vacuum is applied. The aqueous solution is poured onto a silica gel reverse phase column and the desired carbapenem eluted with 5% acetonitrile-water. The appropriate fractions are combined and lyophilized to give the title compound as an amorphous solid (205 mg, 31%).

IR (nujol) v? _max : 1750 (st, β-lactam C = O), 1590 cm " ¹ (st,

C = O) ¹ H NMR (D ₂ O (J: 4.25-3.95 (2H, m, H-1 ', H-5), 3.70-3.40

(6H, m, CH ₂ N ⁺ ), 3.55 (1H, dd, J = 6.1 Hz, J = 2.6 Hz,

H-6), 3.08 (3H, s, CH ₃ N ^+), 3.25 to 2.75 (8H,

H-4) and 1.24 ppm (3H, d, J = 6.4 Hz, CH ₃ ) UV (H ₂ O; c 0.062) _aY 5 299 {t 10962)

1.2

= 17.7 h (0.1 M pH 7 phosphate buffer, 37 ° C).

B e i sp e e 1 17

Preparation of (5R, 6S) -3- [2- (i-Methylmorpholino) ethylthio] -6 - [(R) -1-hydroxyethyl] -7-oxo-1-azabicyclo [3.2.0] - hept -2-ene-2-carboxylate

A. 1-Methyl-1- (2-mercaptoethyl) -morpholineum trifluoro methanesulfonate

Λ / -Λ

+ CF ₃ SO ₃ H + A -> HSCH ₂ CH ₂ NO

CF ₃ SO ₃

10

To N-methylmorpholine (3.29 ml, 0.030 mol) is added dropwise at 10 ⁰ C dropwise trifluoromethanesulfonic acid (1.327 ml, 0.015 mol) and then ethylene sulfide (0.89 ml, 0.015 mol), the resulting yellow-brown solution heated for 18 h N ₂ on the oil bath at 50 to 60 ⁰ C, then removes volatile material in vacuo and the remaining oil in 10 ml of water. The aqueous solution is washed with diethyl ether (3 × 5 ml), then the residual organic solvent is removed in vacuo and the resulting aqueous solution is applied to a C ₁ o-reversed-phase column which is treated with water, then 5% acetonitrile - ^ O and finally Λ0% acetonitrile HpO eluted. Evaporation of the relevant fractions gives a white solid which is dried in vacuo (PpOc) to give the product (1.92 g,

IR (KBr) \? "'. 2560 (-SH) cm " ¹ λ max

¹ H-NMR (dg-acetone) δ: 4.25-3.6 (m, 8H), 3.49 (s, 3H, N-Me), 3.35-2.7 (m, 5H).

B. p-Nitrobenzyl- (5R, 6S) -3- [2- (1-methylmorpholino) -ethylthio] -6 - [(R) -1-hydroxyethyl] -7-oxo-1-azabicyclo [3.2.0] hept-2-ene-2-carboxylate-diphenylphosphate

35

CO ₂ PNB

To a solution of p-nitrobenzyl- (5R, 6S) -6 - [(R) -1-hydroxyethyl] -3,7-dioxo-1-azabicyclo [3.2.0] heptane-2-carboxylate (0.348 g, 1 , 0 mmol) in 25 ml of dry acetonitrile are added dropwise diisopropylethylamine (0.191 ml, 1.1 mmol) and then diphenylchlorophosphate (0.228 ml, 1.1 mmol) at ^{0 0} C under

20 25

gives after 1stün-

Stirring at ⁰ ° C diisopropylethylamine (0.266 mL, 1.3 mmol) to the resulting eno! phosphate and then 1-methyl-1- (2-mercaptoethyl) morpholinium trifluoromethanesulfonate (0.373 g, 1.2 mmol). The reaction mixture is stirred for 1.5 h at room temperature, concentrated in vacuo, the remaining material 'in water and gives on a C ^ g reversed-phase column .. elution with water, then 20% acetonitrile-H ₂ 0 and finally 30% acetonitrile -H ₂ O and subsequent lyophilization of the relevant fractions gives the product (0.360 g, 40%) as an amorphous solid. IR (film): 3300 (-0H), 1770 (β-lactam CO), 1700 (-CO ₂ PNB)

cm"

H-NMR (dg-acetone): 8.25> 7.80 (ABq, J = 8.6 Hz, 4H, arom.), 7.4-6.8 (m, 1OH, diphenyl phosphate), 5, 56, 5.27 (ABq, J = 14.2Hz, 2H, benzyl.), 4.42 (d of t, J = 9.2Hz, J '= 2.7Hz, 1H, H-5) , 4.1-2.7 (m, 17H), 3.40 (s, 3H, N-Me), 1.22 (d, J = 6.2Hz, 3H, -CHMe).

C. (5R, 6S) -3- [2- (i-Methylmorpholino) ethylthio] -6 - [(R) -1-hydroxyethyl] -7-oxo-1-azabicyclo [3.2.0] hept-2 en- 2 ^ -carboxylate

To a solution of p-nitrobenzyl- (5R, 6S) -3- [2- (1-methylmorpholino) -ethylthio] -6 - [(R) -1-hydroxyethyl] -7-oxo-1-azabicyclo [3.2. 0] hept-2-ene-2-carboxylate diphenylphosphate (0.360 g, 0.49 mmol) in 13 ml of phosphate buffer (0.05 M pH 7 * 4) is added 0.36 g of 1096 palladium on carbon , 20 ml of tetrahydrofuran and 20 ml of diethyl ether and hydrogenated de this mixture (Parr) for 1 h under a pressure of 2.2 bar (32 psi). The mixture is filtered through Celite, the filter pad is washed with water and diethyl ether, separating the

aqueous phase and adjust the pH to 7.0 with additional pH 7.4 phosphate buffer. After removing residual organic solvents in vacuo Iieher adding the aqueous solution to a reverse phase column C ^ _Q. Elution with water and lyophilization of the relevant fractions gives 0.130 g of an amorphous solid. This material is purified again by reverse phase HPLC to give the pure product (0.058 g, 34%) as an amorphous solid.

Max

3420 (br, OH), 1750 (β-lactam CO),

-1

IR (KBr) \>

1590 (-CO ₂ ") cm" ¹ H-NMR (D ₂ O) S : 4.35-2.77 (m, 17H), 3.18 (s, 3H, N-Me),

1.23 (d, J = 6.3 Hz, 3H, CH Me) UV (H ₂ 0) \ _max: 300 (£ .6344) mn Τ _{i / 2} (pH 7.4, .36,8 ⁰ C) = 18.5 h.

example

18

Preparation of (5R, 6S) -3- [2- (1,4-dimethyl-1-piperazineum) ethylthio] -6- [i- (R) -hydroxyethyl] -7-oxo-1-azabicyclo Γ 3.2 .0] hept-2-ene-2-carboxylate

COO

A. 1- (2-acetylthioethyl) -1,4-dimethylpiperazinium bromide

+ N N

· Acetone, 5O ⁰ C

A solution of 2-bromoethylthiol acetate (2.20 g, 0.012 mol) [sB Hansen, Acta Chem. Scand. J., 537-40 (1957)] and 1,4-dimethylpiperazine (1.95 mL, 0.014 mol) in 4 mL acetone is stirred for 65 h at 50 ⁰ C. After cooling to 25 ⁰ C Decant the liquid phase from the gummy material which was triturated twice with diethyl ether to give a hygroscopic, yellowish powder (3.2 g, 90? 0 ,

IR (nujol) γ? _Y : 1685 (C = O of the thioester) cm ^-1

¹ H-NMR (D ₂ O) S : 2.37, 2.39 (2s, 6H,

CH ₃ CO, "^ N-CH ₃ ), 3.18 (s, 3H 15

B. 1,4-dimethyl-1- (2-mercaptoethyl) -piperazineumbromidhydο chloride

J, Br

or ,, ^ ₁ S- ^^ S- t i '/ Bl, HCl

A solution of 1- (2-acetylthioethyl) -1,4-dimethylpiperazinium bromide (1.1 g, 3.7 mmol) in 6N hydrochloric acid (4 ml) is heated for 1 h at 8O ⁰ C under an atmosphere stickst off and concentrate the Solution under reduced pressure to give a white powder (0.41 g, 38%).

140 1

1, "λ / 3Γ- ^Η

¹ H-KMR (DMSO-d ₆ ) 6 : 2.90 (s, N ⁺ ), 3.26 (s,

Analysis: for

Calculated: C 31.03% H 7.16% N 9.05% S 10.35% Found: 31.62 7.46 9.19 10.19

'-

C. (5R, 6S) -p-nitrobenzyl-3- [2- (i, 4-dimethyl-i-piperazinilam) -ethylthio] -6- [i- (R) -hydroxyethyl] -7-oxo-1 - azabicvcIo Γ3.2.01hept-2-en-2-carboxylate-diphenyl phosphate

OH ... J-J

Y ^ o

, 0 2) ClP (OPh)

.N

COOPNB

To a cold ⁽⁰ ° C) solution of (5R, 6S) -p-nitrobenzyl-6- [1- (R) -hydroxyethyl] -3,7-dioxo-i-azabicyclo [3.2.0] - heptan-2 - (R) -carboxylate (0.465 g, 1.33 mmol) in 2 mL of acetonitrile (stored under a nitrogen atmosphere) are added diisopropylethylamine (0.278 mL, 1.59 mmol) and diphenylchlorophosphate (0.33 mL, 1.59 mmol). , the reaction mixture is stirred for 30 min and treated with a suspension

Reaction of 1 _f 4-dimethyl-1- (2-mercaptoethyl) piperazinium bromide hydrochloride (0.40 g, 1.37 mmol) in 3 mL acetonitrile-1 mL water mixture and diisopropylethylamine (0.278 mL, 1.59 mmol ). After stirring for 18 h at 5 ⁰ C 15 ml of cold water to the mixture and the resulting solution was chromatographed on a PrepPak-500 C ₁₈ (Waters Associates) column (2.5 x 7.5 cm) to give 25-35 % Acetonitrile in water used as eluent. After lyophilization, a yellowish powder (0.50 g, 50%) is obtained.

IR (KBr) ι? -. 1765 (C = O from β-lactam), 1690 (C = O from PNB ester), 1585 (phenyl), 1512 (NO ₂ ), 875

15 (NO ₂ ) cm " ¹

¹ H-NMR (DMSO-dg) 6 : 1.16, 1.18 (2d, J = 6.1 Hz, 3H, CH, CHOH)

2.44 (s, N-CH ₃ ), 3.14 (s,

'- CH, N "^), 5.31 (d,

J = 6Hz, OH), 5.39 (center of ABq, J = 13Hz, CH ₂ of PNB), 6.6-7.4 (m, 1OH, phenyl of phosphate), 7.71 (d, J = 8.8 Hz, 2H, Ho of PNB), 8.26 (d, J = 8.8 Hz, Hm of PNB).

D. (5R, 6S) -3- [2- (i, 4-dimethyl-1-piperazinium) -ethylthio] -6- [1- (R) -hydroxyethyl] -7-oxo-1-azabicyclo [3 x 2.0 ] hept- 2-ene-2-carboxylate

-, (PhO)

10%, Pd / C ^ THF, ether;

buffer

: OOPNB

, <f

COO

To a solution of (5R, 6S) -p-nitrobenzyl-3- [2- (1,4-dimethy1-1-piperazinium) ethylthio] -6- [i- (R) -hydroxyethyl] -7-oxo Add 1-azabicyclo [3.210] hept-2-ene-2-carboxylate-diphenylphosphate (0.47 g, 0.623 mmol) in 25 mL of moist tetrahydrofuran, diethyl ether (25 mL), potassium phosphate monobasic sodium hydroxide buffer (13 mL , pH 7.22) and 10% palladium on carbon (0.47 g), hydrogenates the resulting mixture for 1 h at 23 ° C under a pressure of 2.76 bar (40 psi), separates the two layers, and Extract the organic layer with water (2 χ 7 ml). The aqueous layers are combined, filtered through a celite pad, washed with diethyl ether (2 × 15 ml) and chromatographed on a PrePak-500 / C ₁₈ (Waters Associates) column (2.5 × 9.5 cm) with water as eluent to give after lyophilization 0.097 g (43%) of product

IR (KBr) _v _ J: 3ΟΟΟ-37ΟΟ (OH), 1750 (C = O of ß-lactam), 1585 (carboxylate) cm "

¹ H-NMR (D ₂ O) S x 1.24 (d, J = 6.4 Hz, 3H, CH ₃ CHOH), 2.33

- \ -ν / -CH-

(s, 3H, ^ N-CH _3), 3.15 (s, _ / ^ _

(m, H-5, CH ₃ CHOH) UV (H ₂ O) A _max : 296 (£ 9,476)

[A] jp. = 61.1 ° (C 0.26, H ₂ O)

ΐ, 1 = 12.4 h (measured at a concentration of

10 ~ ⁴ M in pH 7.4 phosphate buffer at 36, S ⁰ C). 30

Example 19

Preparation of (5R, 6S) -3- [2- (N-methylthiomorpholinium-5-oxide) -ethylthio] -6- [i- (R) -hydroxyethyl] -7-oxo-1-azabicyclo [3.2. 0] hept-2-ene-2-carboxylate

mCPBA

To a cold (-1O ⁰ C) solution of (5R, 6S) -3- [2- (N-methylthiomorpholinium) ethylthio] -6- [1- (R) -hydroxyethyl] -7-OXO-1 -azabicyclo [3.2.0] hept-2-ene-2-carboxylate (608 mg, 1.65 mmol) in a 1: 1 mixture of acetonitrile-water (9 ml) is added in small portions over 1 h.

Chloroperbenzoic acid (334.8 mg, 1.65 mmol), then dilute the mixture with 15 mL of water, wash with diethyl ether (3 X 15 mL), treat the aqueous phase in vacuo, and pass over a reverse phase silica gel column (H ₂ O). in which a solid is obtained which consists of a mixture

consists of connections. This mixture is separated by reverse phase HPLC to give fraction A (52.4 mg, yield 12%) and fraction B (23.6 mg;

Yield 6%) as diastereoisomers of the title compound.

Fraction A ·.

IR (Nujol) \? : 1750 (st, β-lactam C = O), 1580 cm ^-1 (st, C = O)

¹ H-NMR (D ₂ O) S : 4.26-2.91 (2OH, m, H-4, H-5, H-6, H-1 ^! , CH ₂ S, CH ₂ SO, CH ₃ -N ⁺ and CH ₂ N ⁺ ) and 1.24 ppm 3H, d, J = 6.4 Hz, CH ₃ ) UV (H ₂ O, c 0.06) * _max : 302 (£ 10425) t ₁ / ₂ = 12 h (0.065 M, pH 7.4 phosphate buffer, 37 ^° C.

Fraction B

IR (Nujol) v> _v : 1750 (st, β-lactam C = O) and 1585 cm ^-1

(st, C = O)

¹ H-NMR (D ₂ O) 6 x 3.86-2.90 (1H, m, H-4, H-5, H-6, H-1 », CH ₂ S, CH ₂ SO, CH ₂ N ^+), 3.25 (3H, s, CH ₃ N ⁺⁾ and 1.24 ppm (3H, d, J = 6.4 Hz, CH ₃₎ 'UV _(H2 O, c 0.05) A _max : 299 (ε 6517)

^ 1/2 ^{= 10} > ^{75 h} (° » ^o6 5 M, pH 7.4 buffer solution, 37 ° C). 25

Example 20

Preparation of (5R, 6S) -3- [2- (1,4,4-trimethyltetrayl-1-piperazinium) ethylthio] -6- [iR-hydroxyethyl] -7-oxo-1-azabicyclo [3 «2.0 ~) hept-2-en-2-carboxylate chloride 30

A. 1- (2-acetylthioethyl) -1,4,4-trimethylpiperazinium

bromide .jodid

10

Kej

N + N v +, Br, J:

A suspension of 1- (2-acetylthioethyl) -1,4-dimethylpiperazine-bromide (1.48 g, 5.0 mmol) in 10 mL of isopropyl alcohol is treated with methyl iodide (0.373 mL, 6.0 mmol) and heated 3Ph at 55 to 60 ⁰ C. the solvents "medium is at evaporated. reduced pressure, the residue is triturated in hexane, filtered the solid (1.85 g). the Peststoff is dissolved in 8 ml of hot water, the solution diluted to the Turbidity with acetone (70-80 ml) and performs two consecutive crystallizations to give 1.5 g (68%) of the title compound, mp 220-225 ° C (dec.)

IR (KBr) v>: 1692 cm ^-1 (C = O)

a IQaX

¹ H-NMR (D ₂ O) S-. 2.40 (s, 3H, CH ₃ COO), 3.37 (s, N-CH _3), 3.39 (s, N-CH _3), 3.99 (s).

^m ( ^H 2 ^{O) A} max ^{: 22β} (Analysis: for C ₁₁ H ₂ ^ N

Calculated: C 30.08% H 5.51% N 6.38% Found: 30.48 5.53 6.86.

35

B. 1- (2-Mercaptoethyl) -1,4,4-trimethylpiperazinium bis-chloride

HGl 6N permutite

S-1 Cl

2Cl "

A mixture of 1- (2-Acetylthioethyi) -1, 4,4-trimethylpiperaziniumbromid iodide (1.84 g, 4.19 mmol) and 6N hydrochloric acid (15 ml) is heated for 2.5 h under a nitrogen atmosphere at 57 ⁰ C. The solution is concentrated to dryness under reduced pressure, the solid is suspended in 10 ml of water, and the well-stirred mixture is treated with permutite S-1Cl "until a solution is obtained The solution is poured onto a permutite S-1Cl Column (1.2 x 60 cm), which is eluted with water (1.5 ml / min), combine the appropriate fractions and lyophilized to give a white powder (0.93 g, 85%), m.p. 190 to 191 ⁰ C

IR (Nuol) v ^ _max : 2460 (SH).

" ¹ H NMR (O ₂ O) S ', 3.4 Cs, N-CH ₃ ), 3.45 (s, N-CH ₃ ), 4.07

(s) Analysis: for C ₉ H ₂₂ N ₂ SCl ₂ .0.75H ₂ O

Calculated: C 39.34% H 8.62% N 10.20% S 11.67% Found: 39.48 8.39 10.55 11.15.

C. (5R, 6S) -p-nitrobenzyl-3- [2- (i, 4,4-trimethyl-1-piperazinium) -ethylthio] -6- (1R-hydroxyethyl] -7-oxo-1-azabi cyclo Γ3.2.0lhept-2-ene-2-carboxylate-bis-chloride

1) NEt (iPr) ₂

2) ClP (OPh) ₂

COOPNB

3) NEt (iPr) _2v

Permutit S-1 Cl "

2Cl "

COOPNB

2Cl

To a cold (5 ⁰ C) solution of (5R, 6S) -p-nitrobenzyl-6- [iR-hydro xyethyl] -3,7-dioxo-1-azabicyclo [3 · 2.0] heptane-2R-car'boxylat (0.94 g, 2.7 mmol) in 3 ml of acetonitrile (kept under nitrogen atmosphere) are added dropwise diisopropylethylamine (0.557 ml, 3.2 mmol) and diphenylchlorophosphate (0.663 ml, 3.2 mmol), stirring the reaction mixture 30th "at 5 ⁰ C and treated with diisopropylethylamine (0.599 ml, 3.44 mmol) and an aqueous solution (4 ml) of 1- (2-mercaptoethyl) -1,4,4-trimethylpiperazinium-bischlorid (0.90 g , 3.44 mmol) After 1.25 h, diisopropylethylamine (0.1 ml, 0.57 mmol) is added and the mixture is stirred for a further 2 h. A portion of the acetonitrile is removed under reduced pressure and the resulting product is chromatographed. red mixture on a PrePak-500 / CQ (Waters Associates) column using 25-75% acetonitrile in water as the eluent to give, after lyophilization, a yellowish powder (1.4 g) The powder is solubilized in water and added the L solution which is used in a Permutit S-1 Cl ~ column (1.2 χ 58 cm), water as eluent. Lyophilization of the appropriate fractions results in 1.17 g of a. Powder which is in turn purified on a column of PrePak-5Ö0 / C _eighteenth Mari "lyophilizes the appropriate fractions

10

15

and receives a yellowish powder (0.80 g, 53%). IR (KBr) v? _max : 3400 (br, OH), 1770 (C = O of β-lactam), 1690 (C = O of the PNB ester), 1605 (aromat.), 1515

(NO ₂ ), 1345 (NO ₂ ) cm ^{-1 1} H-NMR (O ₂ O) S: 1.26 (d, J = 6.3 Hz, 3H, CH ₃ CHOH), 3.39 (s, NCH ₃ ), 4.00 (s), 5.37 (brs, CH ₂ of PNB), 7.60 (d, J = 8.6 Hz, 2H, Ho of PNB), 8.20 (d, J = 8.7 Hz, 2H, Hm from PNB)

UV (H ₂ O) \ _max : 276 (£ 12094), 306 (t 10752) Analysis: for C ₂₅ H ₃₆ N ^ OgSCl ₂ .3H ₂ O

Calculated: C 46.515a H 6.56% N 8.68 # S 4.97 ^ Cl 10.9890 Found: 46.31 6.18 8.57 5.36 11.37.

D. (5R, 6S) -3- [2- (1,4,4-trimethyl-1-piperazinium) -ethylthio] -6- (1R-hydroxyethyl) -7-0x0-1-azabicyclo [3.2.0] - hept-2-ene-2-carboxylate chloride

COOPNB

30

Cl

35

A mixture of (5R, 6S) -p-nitrobenzyl-3- [2- (1,4,4-trimethyl-1-piperazinium) -ethylthio] -6- (iR-hydroxyethyl) -7-OXO-1-azabicyclo [3.2.0] hept-2-ene-2-carboxylate bis-chloride (0.40 g, 0.68 mmol) * phosphate buffer (30 mL, 0.05 M, pH 7.0), tetrahydrofuran (10 mL ), Ether (30 ml) and 10% Pd / C (0.40 g) are hydrogenated for 1 h at 23 ° C and a pressure of 2.4 bar (35 psi). Separate the two phases, extract the organic phase with water (10 ml), filter the aqueous phase through a celite pad, wash with 10 ml of ether, concentrate in vacuo to 10 ml and chromatograph on a PrepPak 500 / Cg column (2.2 χ 11 cm) (with water as eluent). After lyophilization, 70 mg (25%) are obtained.

IR (KBr); _mo : 3400 (br, OH), 1755 (C = O of β-lactam),

1585 (carboxylate) cm ¹ H-NMR (D ₂ O) S : 1.24 (3H, d, J = 6.3 Hz, CH ₃ CHOH), 3.36 (s, NCH ₃ ), 3.98 ( s) UV (H ₂ O) A _max : 296 (£ 7,987)

[αί ³ = 35.9 ° (c 0.30, H ₂ O)

° i / 2 ⁼ ^ »® k (s ^emessen at a concentration of 10 ~ ⁴ M in pH 7.4 phosphate buffer at 36.8 ⁰ C).

Claims (6)

invention claim 1. . Process for the preparation of carbapenem derivatives h of general formula I. , 8 H , 14 _ COOR ⁴ wherein R is a hydrogen atom and R represents a hydrogen atom or represents the following substituted and unsubstituted radicals: an alkyl, alkenyl and alkynyl group having 1 to 10 carbon atoms; a cycloalkyl and cycloalkylalkyl group having 3 to 6 carbon atoms in the cycloalkyl ring and 1 to 6 carbon atoms in the alkyl moieties; a phenyl group; an aralkyl, aralkenyl and aralkinyl group, wherein the aryl moiety is a phenyl group and the aliphatic moiety is 1 to 6 A heteroaryl, heteroaralkyl, heterocyclyl and heterocyclylalkyl group, wherein the heteroatom or heteroatoms in the above-mentioned heterocyclic units are selected from 1 to 4 oxygen, nitrogen and sulfur atoms and the alkyl moieties associated with the heterocyclic moieties 1 have up to 6 carbon atoms; where the substituent or the substituents of the abovementioned radicals is one of the following groups: a C 1-4 alkyl group, which is optionally substituted by amino, halogen, hydroxy or carboxyl, a halogen atom, 0 0 NR ³ -OR ³ , -OCNR ³ R ⁴ , -CNR ³ R ⁴ , -NR ³ R ⁴ , - NR ³ R ⁴ 0 0 -SO ₂ NR ³ R ⁴ , -NHCNR ³ R ⁴ , R ³ CNR ⁴ -, -CO ₂ R ³ , = 0, 0 0 0 -OCR ^, -SR ", -SR *, -SR *, -CN, -ΝΝ, -OSO ^, η DD -OSO ₉ R ³ , -NR ³ SOpR ⁴ , -NR ³ C = NR ⁴ , -NR ³ COpR ⁴ or pll, -HIl Ο -IYII, - m \. OUpI 15 NO ₂ ,
wherein in the abovementioned substituents the groups • x A Rr and R independently represent a water-20. atom; an alkyl, alkenyl and alkynyl group having 1 to 10 carbon atoms; a cycloalkyl, cycloalkylalkyl and alkylcycloalkyl group having 3 to 6 carbon atoms in the cycloalkyl ring and having 1 to 6 carbon atoms in the alkyl moieties; a phenyl group; an aralkyl, aralkenyl and etra-kynyl group wherein the aryl moiety is a phenyl group and the aliphatic moiety has from 1 to 6 carbon atoms; or a heteroaryl, heteroaralkyl, heterocyclyl and heterocyclylalkyl group, wherein the heteroatom or heteroatoms in the above-mentioned heterocyclic units are selected from 1 to 4 oxygen, nitrogen or sulfur atoms and the alkyl moieties associated with said heterocyclic moieties have 1 to 6 carbon atoms have, stand, or 35 3 4
R ^ and R together with the nitrogen atom, to which is bonded to at least one of these radicals, form a 5- or 6-membered, nitrogen-containing, heterocyclic ring; R ^ is defined as the group R ^ but can not be hydrogen;
or in which R and R together are a C2_ ₁₀ alkylidene or substituted by a hydroxy group mean C2_ ₁₀ alkylidene; A represents a cyclopentylene, cyclohexylene or a C ₂ _g-alkylene group, which is optionally substituted by one or more C ₁ ^ -alkyl groups; 2 "~ R is a hydrogen atom, an anionic charge or a common, easily removable carboxyl-protecting 2, provided that when R represents a hydrogen atom or a protecting group, also .20 a counteranion is present, and 14 R is a quaternized, nitrogen-containing, aromatic or non-aromatic heterocycle bonded to A via a ring nitrogen atom, forming a quaternary ammonium group, or a pharmaceutically acceptable salt thereof, characterized in that an intermediate compound of the general formula IV · _O 8 H ,1. R ' I 2 ' COOR IV Λ R in which R and R have the meanings given above 2 »
R is a common, readily removable carboxyl protecting group and L is a common leaving group,
with a thio compound of the general formula VII HS-A-R VII , 14 wherein A and R have the meanings given above and Χ ^{θ represents} a counter anion, in an inert solvent and in the presence of a base to a carbapenem compound of general formula I ^f , 8 H , Θ I ¹ wherein R ¹ , R ⁸ , R ² ', A, r' have 25 meanings, a
and Χ ^σ the above and, if desired, the carboxy1-protecting group 2 ' R to obtain the corresponding deblocked compound of general formula I or a pharmaceutically acceptable salt thereof. 2. Method according to item 1, characterized by in that the base used is a non-nucleophilic, tertiary amine or a Trl (C._λ) alkylamine. 3. The method according to item 2, characterized'dadurch, that is used as the base, a non-nucleophilic tertiary amine. 4. The method according to item -1, 2 or 3, characterized in that one carries out the reaction at a temperature of -15 ° C to room temperature. 5. The method according to item 4, characterized in that the reaction at a temperature of about -15 ° C to +15 ⁰ C performed. 6. A process according to any one of items 1 to 5, characterized in that acetonitrile, a mixture of acetonitrile, dimethylformamide and tetrahydrofuran, a mixture of tetrahydrofuran and water, a mixture of acetonitrile and water and acetone are used as the inert solvent. - 7. Procedure for the preparation of a quaternary Aminthiol compound of the general formula VIL HS-A-R ¹⁴ Χ ^θ VII, "' wherein A is cyclopentylene, cyclohexylene or - R ¹⁰ R ¹² X yy 10 11 12 '1 "i stands;' in which R, R, R and R 1 are independently of one another 5-of-one-tof a-or-a-C 1-6-alkyl group.