CH633800A5 - Process for the preparation of guanidines - Google Patents

Abstract

Guanidines of the general formula II <IMAGE> in which the substituents are defined in Claim 1, and their pharmaceutically acceptable salts are obtained by treating thienamycin (I) with a urea derivative, with intermediate protection of the oxygen functions and/or the carboxyl function, and converting compounds which may be obtained into their pharmaceutically acceptable salts. The novel compounds which can be prepared according to the invention are novel useful antibiotics which as a rule are active against various gram-positive and gram-negative bacteria and which are therefore used in human and veterinary medicine.

Description

The invention relates to a process for the preparation of guanidines. Such compounds which can be prepared according to the invention and their pharmaceutically acceptable salt derivatives are usually suitable as antibiotics.

Thienamycin is described and claimed in US Pat. No. 3,950,357. This patent is hereby incorporated into the disclosure because thienamycin is used as the starting compound in the process according to the invention. The anti-bioticum thienamycin and its preparation are described in DT-OS 25 52 638. It is available, for example, by cultivating the microorganism Streptomyces cattleya (MA-4297), a culture of which is in the culture collection of the Northern Regional Research Laboratories, Northern Utilization Research and Development Division, Agricultural Research Service, US Department of Agriculture, Peoria, Illinois, V.St.A and is designated with NRRL 8057.

Thienamycin has the following structural formula:

with intermediate protection of the oxygen functions and / or the carboxyl function, with a compound of the formula

X "R °

I.

R, R2N-C = N-R1

Oh

/> 0

-n-

sch2CH2NH2 cooh

3rd

633 800

The substituted N-methylene thienamycin derivatives according to the invention can be characterized by the following structural formula:

II

sch2ch2n = c-x cooh y which, depending on the basicity of the amino nitrogen (a function of the identity of the methylene substituents X and Y), can also be represented as an equivalent inner salt:

Oh

/? 0

.CO (ß

h sch? ch -, _ k = ç-x

II '

20th

which is a recognized form of a simple resonance structure, for example when Y is -NR'R2 and X is R:

Oh

©

K ~ ^ 12

sch2ch2k - = ^ - c —kr r

II "

-Q

oy

For the sake of simplicity, the connections that can be produced according to the invention can be represented by the symbol

OH

Th-4-N = C-X I

Y

L-COOH

35

II

40

are reproduced, in which "Th" represents the bicyclic nucleus of thienamycin and its hydroxyl-, amino- and carboxyl-functional groups are shown and in which X and 45 Y are selected independently of one another from the group hydrogen, -R, -OR, -SR and - NR'R2; and wherein R1 and R2 are selected independently of one another and R, hydrogen, nitro, hydroxyl, alkoxyl having 1 to 6 carbon atoms, amino, mono-, di- and trialkylamino, in which the alkyl group in each case comprises 1 to 6 carbon atoms; R1 and R2 can also be linked together to form a substituted or unsubstituted mono- or bicyclic heteroaryl or heterocyclyl containing (together with the nitrogen atom to which they are attached) 4 to 10 atoms, one of which additionally has 55 or more Heteroatoms can be selected from the group consisting of oxygen, sulfur or nitrogen; R 'and R2 are substituted or unsubstituted: cyano; Carbamoyl;

Carboxyl; Alkoxycarbonyl and alkyl of 1 to about 10 carbon atoms; Alkenyl of 2 to about 10 carbon atoms; 60 alkynyl with 2 to about 10 carbon atoms; Cycloalkyl of 3 to 10 carbon atoms; Cycloalkylalkyl and cycloalkylalkenyl having 4 to 12 carbon atoms; Cycloalkenyl, cycloalkenylalkenyl and cycloalkenylalkyl each having 3 to 10.4 to 12 and 4 to 12 carbon atoms; Aryl of 6 to 10 carbon atoms, aralkyl, aralkenyl and aralkynyl of 7 to 16 carbon atoms; mono- and bicyclic heteroaryl and heteroalkyl, which typically contains 4 to 10 ring atoms, one or more of which can be a heteroatom from the group consisting of oxygen, sulfur or nitrogen, and in which the alkyl radical of the heteroalkyl radical contains 1 to about 6 carbon atoms ; mono- and bicyclic heterocyclyl and heterocyclylalkyl, which typically contains 4 to 10 ring atoms, one or more of which can be a heteroatom from the group selected from oxygen, sulfur or nitrogen and in which the alkyl radical of the heterocyclylalkyl radical 1 b's contains about 6 carbon atoms; the aforementioned substituent or the substituents on R, R1, R2 or on the ring formed by the connection of R1 and R2 can be selected from the group: halogen, such as chlorine, bromine, iodine and fluorine ; Azido; Alkyl of 1 to 4 carbon atoms; Thio; Sulpho; Phosphono; Cyanothio (-SCN); Nitro; Cyano; Amino; Hydrazino; mono-, di- and trialkyl-substituted amino and hydrazino, wherein the alkyl has 1 to 6 carbon atoms; Hydroxyl;

Alkoxyl of 1 to 6 carbon atoms; Alkylthio with 1 to 6 carbon atoms; Carboxyl; Oxo; Alkoxycarbonyl having 1 to 6 carbon atoms in the alkoxyl group; Acyloxy containing 2 to 10 carbon atoms; Carbamoyl and mono- and dialkylcarbamoyl, wherein the alkyl group has 1 to 4 carbon atoms.

There is generally a constant need for new antibiotics. Unfortunately, there is usually no static effectiveness for a given antibiotic because continued use on a broad scale of such an antibiotic selectively increases the formation of resistant strains of pathogens. In addition, the known antibiotics have the disadvantage, for example, that they are only effective against certain types of microorganisms. That is why research is usually continued on new antibiotics.

It has now been found, unexpectedly, that the compounds which can be prepared according to the invention are usually broad-spectrum antibiotics which are useful for both animal and human therapy and in inanimate systems.

It is therefore an object of the present invention, for example, to provide a new class of antibiotics which have the basic core structure of thienamycin (I), but which are characterized in that they are substituted N-methylene derivatives thereof. These antibiotics are usually active against a wide range of pathogens, for which both gram-positive bacteria are representative, such as S. aureus, Strep. pyogenes and B. subtilis, and gram-negative bacteria such as E. coli, Proteus morganii, Klebsiella, Serratia and Pseudomonas are representative. Another object of the invention is, for example, to provide a chemical process for the production of such antibiotics and their non-toxic pharmaceutically acceptable salts.

The guanidines of the formula II which can be prepared according to the invention, in which

Y = -NR'R2 and X = -NR'R2 is:

can be abbreviated by the formula - OR3

Th-1- N = C-NR'R2 I

NR'R2 LCOX'R3 '

in which all symbols have the meaning given above.

633 800

4th

Typical examples of such guanidine embodiments (the substituent and the amino group of the thienamycin form the guanidine structure) are:

Guanidine: Y = -NH2, X = -NH2;

N-methylguanidine: Y = -NHCH3, X = -NH2; N, N-dimethylguanidine: Y = -N (CH3) 2, X = -NH2; N, N, N-trimethylguanidine: Y = -N (CH3) 2, X = -NHCH3; N-phenylguanidine: Y = -NH (C6H5), X = -NH2;

Nitroguanidine: Y = -NHN02, X = -NH2;

Aminoguanidine: Y = -NHNH2, X = -NH2.

Other preferred guanidines which can be prepared according to the invention are those in which R1 and R2 are selected independently of one another from the group consisting of: hydrogen; substituted and unsubstituted groups: alkyl of 1 to 6 carbon atoms such as methyl, ethyl, isopropyl, butyl, tert-butyl, N, N-dimethylaminoethyl, 2,2,2-trifluoroethyl, 2-methylthioethyl and the like; Alkenyl of 3 to 6 carbon atoms such as allyl, methallyl, 2-butenyl, l-buten-3-yl and the like; Cycloalkyl, cycloalkylalkyl, cycloalkenyl and cyclo-alkenylalkyl each having 3 to 6.4 to 7.4 to 6 and 4 to 7 carbon atoms, such as cyclopropyl, cyclopentyl, cyclohexyl, cyclopropylmethyl, cyclohexylmethyl, 2-cyclopropenyl, 1,4-cyclohexadienylmethyl, and the like; Aralkyl and aralkenyl with 7 to 10 carbon atoms such as benzyl, p-methoxy-benzyl, p-dimethylaminobenzyl, cinnamyl and the like; monocyclic heteroaralkyl with 5 to 6 ring atoms, one or more of which are selected from oxygen, sulfur and nitrogen, and 1 to 3 carbon atoms in the alkyl radical, such as 2-thienylmethyl, 3-thienylmethyl, 2-furylmethyl, 1-methyl-5 -tetrazolylmethyl and the like; wherein the ring or chain substituent may be chlorine, fluorine, hydroxyl, alkoxyl of 1 to 3 carbon atoms, dialkylamino of 1 to 3 carbon atoms in each alkyl group, alkylthio of 1 to 3 carbon atoms in the definition of R1 and R2;

X 'is oxygen and R3 and R3' are hydrogen.

The compounds which can be prepared according to the invention are prepared in a simple manner from thienamycin (I, as previously indicated). Embodiments in which the secondary alcohol group and / or the carboxyl group are derived can be prepared in a simple manner, either from the corresponding O-, carboxyl- or O- and carboxyl derivative of thienamycin or from II or thienamycin by subsequently carrying out a reaction, to form residues R3 and R3 '(or —X'R3') and combinations thereof. Such starting materials are described, for example, in the following US patent applications, which are to be included hereby: US application serial number 634 006 dated November 21, 1975, which relates to the O-derivatives of thienamycin (ester and ether derivatives of the secondary alcohol group of thienamycin) is aligned and which has the case number 15821YA and which is characterized by the following structural formula:

Oh

«I

SCH ^ CH ^ NR1 3T coch

Ib

5 ér wherein R1 'and R2' are selected from the group consisting of hydrogen and acyl; the term "acyl" is defined as in the patent application given below. 10 Such N-acylthienamycins are, for example, useful starting compounds for the preparation of corresponding substituted pseudoureas and imido ethers and imido thioethers.

US Application Serial Number 634,298 dated November 21, 15 1975 with the internal designation 15818YA, which is directed to carboxyl derivatives of thienamycin with the following structural formula:

OH

cT

_N_

sch2ch2wh2

cox'r3

Ic

25th

US Application Serial Number 634,295 dated November 21, 1975 with the internal designation 15813YA, which is directed to N-acyl and carboxyl derivatives of thienamycin with the following structural formula:

30th

35

oh jf

_K-

11 2 check no

Id cox'r

3 '

US application serial number 634,294 dated November 21, 1975 with the internal name 15822YA, which is directed to N-acyl, O and carboxyl derivatives of thienamycin with 1 of the following structural formula:

, 3rd

OR

45

4th

✓ / nv-sch0ch0nr1 r2

MI

n LLcox'r3

Ie

OR '

sch2ch2sh2

cooh

Thus, the embodiments of the present invention as set forth above with IIa can be made by starting with the corresponding derivatives la, Ib, Ic, Id and Ie, or embodiment IIa can be made by working directly with Thienamycin I (I—> 11) begins and then includes the derivatives to form the 55 groups R3 and / or X'R3 '(II— »IIa), as described in the previously mentioned and hereby incorporated US patent applications fertilize has been described.

With regard to structures Ia, Ib, Ic, Id and Ie, the 60 radicals R3, R3 ', X' and acyl (Rr and R2 ') can be defined as follows:

Identification of the rest -COX 'R3'

In the general description of compounds (IIa) US application Serial Number 634,291 dated November 21, 65, the remainder designated by the group -COX'R3 'means 1975 with the case character 15775YA, which is aimed at, below other -COOH (X 'is oxygen and R3' is what-N-acyl derivatives of thienamycin with the following structural substance) and all residues which are known to be effective pharmaceutical formulas: tacitically acceptable esters, anhydrides (R3 'is acyl) and

5 633 800

Amide residues in bicyclic β-lactam antibiotics, such as in the 2-methylaminoethyl, 2-diethylaminoethyl, 2-acetamidoethyl,

Cephalosporins and penicillins and their core ana- phthalimidomethyl, succinimidomethyl; Alkoxyalkyl,

lied. wherein the alkoxy part 1 to 10 and preferably 1 to 6 carbon

Suitable radicals (R3) include conventional protective or material atoms and have branched, straight-chain or cyclic carboxyl blocking groups. The term “blocking group 5, and in which the alkyl part has 1 to 6 carbon atoms,” as used here, is used in the same way as methoxymethyl, ethoxymethyl, isopropoxymethyl, decyl manner as used in accordance with US Pat. PS 3,697,515, which is included here-oxymethyl, ethoxypropyl, decyloxypentyl, cyclohexyloxyme with respect to the disclosure, uses thyl and the like; Alkanoyloxyalkyl, wherein the alkanoyl is. Pharmaceutically acceptable thienamycin derivatives, the oxy part is straight or branched and 1 to 6 carbon present invention, which fall under this class, will have 10 atoms and the alkyl part has 1 to 6 carbon atoms, as indicated below. Suitable blocking esters include acetoxymethyl, pivaloyloxymethyl, acetoxyethyl, propionyl-thus those as specified in the following list, oxyethyl, acetoxypropyl and the like. Haloalkyl, in which are, which is representative but is in no way an exhaustive halogen, chlorine, bromine, fluorine or iodine and the alkyl part is an enumeration of the possible ester groups in which the chain or branched chain has 1 to 6 carbon atoms, X '= 0 and R3' are given: 15 for example 2,2,2-trichloroethyl, trifluoroethyl, 2-bromopro-

(i) R3 '= CRaRbRc, wherein at least one of Ra, Rb and Rc pyl, diiodomethyl, 2-chloroethyl, 2-bromoethyl and the like; is an electron donor, for example p-methoxyphenyl, alkenyl with 1 to 10 carbon atoms, which is either straight-2,4,6-trimethylphenyl, 9-anthryl, methoxy, CH2SCH3, tetra-chain or branched, for example allyl, 2-propenyl , hydrofur-2-yl, tetrahydropyran-2-yl or fur-2-yl. The remaining 3-butenyl, 4-butenyl, 4-pentenyl, 2-butenyl, 3-pentenyl,

Chen Ra, Rb and Rc groups can be hydrogen or organic 20 3-methyl-3-butenyl, methallyl, 1,4-cyclohexadien-l-yl-methyl see substitution groups. Suitable ester groups of these and the like; Alkynyl with 1 to 10 carbon atoms, that

Kind include a p-methoxybenzyloxycarbonyl and 2,4,6-tri- either straight-chain or branched, for example methylbenzyloxyearbonyl. 3-pentynyl, propargyl, ethynyl, 3-butyn-l-yI and the like;

(ii) R3 '= CRaRbRc, wherein at least one of the Ra, Rb and alkanoyl, which can be either straight-chain or branched, Rc groups is an electron-attracting group, for example having 1 to 10 carbon atoms, such as pivaloyl, acetyl, propionyl such as benzoyl, p-nitrophenyl, 4-pyridyl, trichloromethyl, tri- and the like; Aralkyl or heteroalkyl, in which the alkyl has 1 bromomethyl, iodomethyl, cyanomethyl, ethoxycarbonylme to 3 carbon atoms, and hetero means that 1 to 4 thyl, arylsulphonylmethyl, 2-dimethylsulphoniummethyl, heteroatoms are present, selected from the group be-o-nitrophenyl or cyano. Suitable esters of this type, including O, S or N, such as benzyl, benzhydryl and substitute a benzoylmethoxycarbonyl, p-nitrobenzyloxycarbonyl, 30 iert benzyl, benzhydryl, for example benzyl or benzhy-4-pyridylmethoxycarbonyl, 2,2,2- Trichloroethoxycarbonyl and dryl substituted with 1 to 3 substituents such as benzyl, phenoxy, 2,2,2-tribromoethoxycarbonyl. Halogen, lower alkyl, lower alkanoyloxy with 1 to 5 carbon

(iii) R3 '= CRaRbR °, wherein at least two of the Ra, Rb and substance atoms, lower alkoxy, hydroxy, nitro, blocked Rc radicals are hydrocarbons, such as alkyl, for example carboxy or combinations thereof, for example p-chloro-methyl or ethyl or Aryl, for example phenyl and the 35 benzyl, o-nitrobenzyl, 3,5-dinitrobenzyl, p-methoxybenzyl, residual Ra, Rb and Rc group, if present, hydrogen m-benzoylbenzyl, p-tert-butylbenzyl, m-phenoxybenzyl , means. Suitable esters of this type include tert-butyl-p-benzoylbenzyl, p-nitrobenzyl, 3,5-dichloro-4-hydroxybenzyl, loxycarbonyl, tert-amyloxycarbonyl, diphenylmethoxycarbo-p-methoxycarbonylbenzyl, p-methoxybenzhydryl, p-carboxy- nyl and triphenylmethoxycarbonyl. benzyl, the latter either as a free acid, as an ester

(iv) R3 '= Rd, where Rd is adamantyl, 2-benzyloxyphenyl, 40 or as the sodium salt, 2,4,6-trimethylbenzyl, p-pivaloy-4-methylthiophenyl or tetrahydropyran-2-yl. loxybenzyl, p-tert-butoxycarbonylbenzyl, p-methylbenzyl,

Silyl esters of this category of blocking groups can- p-benzoyloxybenzyl, p-acetoxybenzyl, p-2-ethylhexanoyI-

be easily produced from halosilanes benzyl, p-ethoxycarbonylbenzyl, p-benzoylthiobenzyl, p-benz

or silazanes of the formula: amidobenzyl, o-pivaloyloxybenzyl, m-pivaloyloxybenzyl, p-iso-

R43SiX '; R42SiX'2; R43Si.NR42; R43Si.NH.COR4; 45 propoxybenzyl, p-tert-butoxybenzyl and also the cyclic

R43Si.NH.CO.NH.SiR43; R4NH.CO.NH.SiR43; or R4C analogues thereof, 2,2-dimethyl-5-coumaranmethyl, 5-indanylme-

(OSiR43); HN (SiR43) 2, wherein X 'is halogen, chlorine or bromothyl, p-trimethylsilylbenzyl, 3,5-bis-t-butoxy-4-hydroxybenz-

means and the different groups R4, the same or yl; 2-thienylmethyl, 2-furylmethyl, 3-tert-butyl-5-isothiazole

can be different, hydrogen atoms or alkyl, meaning methyl, 6-pivaloyloxy-3-pyridazinylethyl, 5-phenylthio-1-tetra-

ten, for example methyl, ethyl, n-propyl, isopropyl; Aryl, such as zolylmethyl or the like (the term lower alkyl, for example, phenyl; or aralkyl, for example a benzyl or lower alkoxy in this context a chain with 1

group. means up to 4 carbon atoms); or phthalidyl; or phhe

More generally, pharmaceutically acceptable nylethyl, 2- (p-methylphenyl) ethyl and the arylthioalkylanalo-

Carboxyl derivatives in general those derived from ge, aryloxyalkyl, wherein aryl is preferably a phenyl ring with

Reacting thienamycin or an N-protected thien 55 0 to 3 substituents, preferably 0 or 1 substituents in amyein, such as an N-acylated thienamycin, with alcohols, the ortho or para position and the alkyl 1 to 6 carbon phenols , Mercaptans, thiophenols, acylating agents and atoms, for example (4-methoxy) phenoxymethyl, pheno-

the like. For example, interesting esters and amides are xymethyl, (4-chloro) -phenoxymethyl, (4-nitro) -phenoxymethyl,

the previously listed starting compounds and endpro- (4-benzyloxy) phenoxymethyl, (4-methyl) phenoxymethyl,

products containing the following groups in the 2-position of 60 (2-methoxy) phenoxymethyl, (l-phenoxy) ethyl, (4-amino) -

Thienamycin nucleus have: -COX'R3 ', where X' is oxygen, phenoxymethyl, (4-methoxy) -phenylthiomethyl, (4-chloro) -

Is sulfur or NR '(R' = H or R3 ') and R3 is alkyl phenylthiomethyl, phenylthioethyl; Aryl, where aryl is phenyl,

means with 1 to 10 carbon atoms, specifically straight-line 5-indanyl or substituted phenyl with 0 to 3 substituents,

or branched, such as methyl, ethyl, tert-butyl, pentyl, decyl, preferably 0 to 1 substituents in the ortho- or para

and the same; Carbonylmethyl, including phenacyl, means 65 position, for example (4-methyl) phenyl, (4-hydr

p-bromophenacyl, p-tert-butylphenacyl, acetoxyacetylmethyl, oxy-phenyl, (4-tert-butyl) -phenyl, p-nitrophenyl, 3,5-dinitro-

Pivaloxyacetylmethyl, carboxymethyl and its alkyl and phenyl or p-carboxyphenyl, the latter either as

Aryl ester, a-carboxy-a-isopropyl; Aminoalkyl including free acid or as sodium salt; Aralkenyl, wherein aryl

633 800

Phenyl or alkenyl having 1 to 6 carbon atoms means such as 3-phenyl-2-propenyl; Aralkoxyalkyl, wherein aralkoxy is benzyloxy and the alkyl has 1 to 3 carbon atoms, such as benzyloxymethyl, (4-nitro) -benzyloxymethyl, (4-chloro) -benzyloxymethyl; Alkylthioalkyl, in which the alkylthio part has 1 to 10 and preferably 1 to 6 carbon atoms, but which can be branched, straight-chain or cyclic and the alkyl part has 1 to 6 carbon atoms, such as methylthioethyl, ethylthioethyl, cyclohexylthiomethyl, decylthiobutyl, methylthiopropyl, isopropylthioethyl, Methylthiobutyl and the like.

In addition to the esters (and thioesters) listed above, the present invention also encompasses, for example, amides, that is, in which X '

R '

is a —N group. Typical examples of such amides are those in which R 'is selected from the group consisting of hydrogen, methyl, ethyl, phenyl, p-methoxyphenyl, benzyl, carboxymethyl, methylthioethyl and heteroaryl; The grouping -COX'R3 'also includes anhydrides in which R3' is acyl, for example benzyloxycarbonyl, ethoxycarbonyl, benzoyl and pivaloyl.

The most preferred -COX'R3 'residues of the present invention are those in which, relative to structure IIa, X' represents oxygen, sulfur or NR '(R' is selected from the group consisting of hydrogen and lower alkyl); and R3 'is selected from the group consisting of: lower alkyl, lower alkenyl such as methallyl, 3-methylbutenyl, 3-butenyl, and the like; Methylthioethyl; Benzyl and substituted benzyl such as p-tert-butylbenzyl, m-phenoxybenzyl, p-pi-valoyloxybenzyl, p-nitrobenzyl and the like; Pivaloyloxy-methyl, 3-phthalidyl and acetoxymethyl, propionyloxymethyl, acetylthiomethyl, pivaloylthiomethyl, allyl, 4-butenyl, 2-butenyl, 3-methyl-2-butenyl, phenacyl, acetoxyacetylmethyl, methoxymethyl, p-acetbenzylbenzylbenzylbenzyl p-iso-propoxybenzyl, 5-indanylmethyl, 5-indanyl, benzyloxymethyl, ethylthioethyl, methylthiopropyl, methoxycarbonyloxymethyl, ethoxycarbonyloxymethyl, dimethylaminoacetoxymethyl, crotonolacton-3-yl and acetamidomethyl.

Identification of R3 (Rr and R2 ')

In a general representation of the present invention, in structure IIa the radical R3 can be hydrogen, except 1.) acyl (in general the group -OR3 is referred to as an ester); or 2.) R3 is selected from alkyl, aryl, aralkyl and the like, so that the group -OR3 can be referred to as an ether. For the ester version (1), R3 is preferably selected from the following definitions for acyl residues (p = 1). In the so-called ether version (2.) of the present invention, R3 is selected, for example, from the same acyl radicals in which the carbonyl group

O X

-C- or generally -C- is omitted (p = 0); so that R3 is selected from the following radicals, in which all symbols are designated below:

Rr and R2 'are normally selected from the above radicals in which p = 1. Therefore, in relation to the

Definition of R3, R1 'and R2' of the acyl radical can be, inter alia, a substituted or unsubstituted aliphatic, aromatic or heterocyclic, araliphatic or heterocyclylaliphatic carboxylic acid radical, a substituted or unsubstituted carbamyl radical or a carbothioic acid radical. A group of acyl residues can be represented by the general formula:

X II

-C-R ",

where X = 0 or S and R "is hydrogen; amino; substituted amino, such as alkyl- and dialkylamino, wherein the alkyl radical comprises 1 to 6 carbon atoms; substituted or unsubstituted radicals, such as: straight-chain or branched-chain alkyl radicals, wherein the alkyl radical Comprises 1 to 6 carbon atoms; mercapto; aryloxy, which typically comprises 6 to 10 carbon atoms; alkenyl or alkynyl groups, which typically comprise 2 to 6 carbon atoms; aryl, such as phenyl; aralkyl, such as benzyl; cycloalkyl, which is typically 3 comprises up to 6 carbon atoms; or a heteroaryl or heteroaralkyl group (mono- or bicyclic), in which the alkyl group typically comprises 1 to 3 carbon atoms and the heterocyclic ring typically contains 4 to 10 atoms and the heteroatom or the heteroatoms are selected from O, N and S; the aforementioned groups can be unsubstituted or they can be substituted by radicals such as OH, SH, SR (R is N Lower alkyl or aryl, such as phenyl), alkyl or alkoxy groups having 1 to about 6 carbon atoms, halogen, such as Cl, Br, F and J, cyano, carboxy, sulfamino, carbamoyl, sulfonyl, azido, amino, substituted amino, such as alkylamino including quaternary Ammonium, in which the alkyl groups have 1 to 6 carbon atoms, haloalkyl, such as trifluoromethyl, carboxyalkyl, carbamoylalkyl, N-substituted carbamoylalkyl, in which the alkyl group of the aforementioned four radicals comprises 1 to about 6 carbon atoms, amidino, guanidino, N-substituted guanidino , Gu-anidino lower alkyl and the like. Typical examples of such acyl groups which can be mentioned here are those in which R "denotes benzyl, p-hydroxybenzyl, 4-amino-4-carboxybutyl, methyl, cyanomethyl, 2-pentenyl, n-amyl, n-heptyl, Ethyl, 3- or 4-nitrobenzyl, phenethyl, ß, ß-diphenylethyl, methyldiphenylmethyl, triphenylmethyl, 2-methoxyphenyl, 2,6-dimethoxyphenyl, 2,4,6-trimethoxyphenyl, 3,5-dimethyl-4 -isoxazolyl, 3-butyl-5-methyl-4-isoxazolyl, 5-methyl-3-phenyl-4-isoxazolyl, 3- (2-chlorophenyl) -5-methyl-4-isoxazolyl, 3- (2,6- Dichlorophenyl) -5-methyl-4-isoxyzolyl, D-4-amino-4-carboxybutyl, D-4N-benzoylamino-4-carboxy-n-butyl, p-aminobenzyl, o-aminobenzyl, m-aminobenzyl, p-dimethylaminobenzyl , (3-pyridyl) methyl, 2-ethoxy-1-naphthyl, 3-carboxy-2-quinoxalinyl, 3- (2,6-dichlorophenyl) -5- (2-furyl) -4-isoxazolyl, 3-phenyl -4-isoxazolyl, 5-methyl-3- (4-guanidinophenyl) -4-isoxazolyl, 4-guanidinomethylphenyl, 4-guanidinomethylbenzyl, 4-guanidinobenzyl, 4-guanidino-phenyl, 2,6-dimethoxy-4 ~ guanidino, o -Sulfobenzyl, p-carb -oxymethylbenzyl, p-carbamoylmethylbenzyl, m-fluorobenzyl, m-bromobenzyl, p-chlorobenzyl, p-methoxybenzyl, 1-naphthyl-methyl, 3-isothiazolylmethyl, 4-isothiazolylmethyl, 5-isothi-azolylmethyl, 4-pyranylthiomethyl -Isoxazolyl-methyl, 4-methoxy-5-isoxazolylmethyl, 4-methyl-5-isoxazolyl-methyl, 1-imidazolylmethyl, 2-benzofuranylmethyl, 2-indolyl-methyl, 2-phenylvinyl, 2-phenylethynyl, 1-aminocyclohexyl, 2 and 3-thienylaminomethyl, 2- (5-nitrofuranyl) vinyl, phenyl, o-methoxyphenyl, o-chlorophenyl, o-phenylphenyl, p-aminomethylbenzyl, l- (5-cyanotriazolyl) methyl, difluoromethyl, dichloromethyl, Dibromomethyl, 1- (3-methylimidazolyl) methyl, 2- or 3- (5-carboxymethylthienyl) methyl, 2- or 3- (4-

6

5

10th

15

20th

25th

30th

35

40

45

50

55

60

633 800

Carbamoylthienyl) methyl, 2- or 3- (5-methylthienyl) methyl, 2- or 3- (methoxythienyl) methyl, 2- or 3- (4-chlorothienyl) methyl, 2- or 3 - (5-Sulfothienyl) methyl, 2- or 3- (5-carboxythienyl) methyl, 3- (l, 2,5-thiadiazolyl) methyl, 3- (4-methoxy-l, 2,5-thiadiazolyI ) -methyl, 2-furylmethyl, 2- (5- 5 nitrofuryl) -methyl, 3-furylmethyl, 2-thienylmethyl, 3-thienylmethyl, tetrazolylmethyl, benzamidinomethyl and cyclo-hexylamidinomethyl.

The acyl group can also be a radical of the formula:

X

10th

-C (CH2) »ZR",

where X is O or S and n is 0 to 4 and Z is oxygen,

15

Is sulfur, carbonyl or nitrogen and R "has the meaning given above. Typical members of the substituent

- (CH2) nZR ",

20th

which should be mentioned here are allylthiomethyl, phenylthio-methyl, butylmercaptomethyl, a-chlorocrotylmercaptomethyl, phenoxymethyl, phenoxyethyl, phenoxybutyl, Pher.oxy-benzyl, diphenoxymethyl, dimethylmethoxyethyl, dimethyl-butoxymethyl, dimethylphenoxymophenyl, 4-guyloxymethylphenyl 4-pyridylthiomethyl, p- (carboxymethyl) phenoxymethyl, p- (carboxymethyl) phenylthiomethyl, 2-thiazolylthiomethyl, p- (sulfo) phenoxymethyl, p- (carboxymethyl) phenylthiomethyl, 2-pyrimidinylthiomethyl, Phenethylthiomethyl, 1- (5,6,7,8-tetrahydronaphthyl) oxomethyl, N-methyl-4-pyridylthio, benzyloxy, methoxy, ethoxy, phenoxy, phenylthio, amino, methylamino, dimethylamino, pyridiniummethyl, tri-methylammoniummethyl , Cyanomethylthiomethyl, trifluoromethylthiomethyl, 4-pyridylethyl, 4-pyridylpropyl, 4-pyridylbutyl, 3-imidazolylethyl, 3-imidazolylpropyl, 3-imidazolylbutyl, 35 1-pyrroloethyl, 1-pyrrolopropyl.

Alternatively, the acyl group can be a radical of the formula

X II

-C-CHR "

I.

R, "

where R "has the meaning given above and R '" is a radical such as amino, hydroxy, azido, carbamoyl, guanidino, amidino, acyloxy, halogen, such as chlorine, fluorine, bromine, iodine, sulfamino, tetrazolyl, Sulfo, carboxy, carbalkoxy, phosphono and the like. Typical members of the substituent

50

-CHR "

R '

55

which should be mentioned here are a-aminobenzyl, a-amino- (2-thienyl) -methyl, a- (methylamino) -benzyl, a-aminomethyl-mercaptopropyl, a-amino-3- or -4-chlorobenzyl, a -Amino-3- or -4-hydroxybenzyl, a-amino-2,4-dichlorobenzyl, a-amino-3,4-dichlorobenzyl, D (-) - a-hydroxybenzyl, a-carboxybenzyl, a-amino- (3-thienyl) methyl, D (-) - a- 60

Amino-3-chloro-4-hydroxybenzyl, a-amino- (cycIohexyl) -methyl, a- (5-tetrazolyl) -benzyl, 2-thienylcarboxylmethyl, 3-thienylcarboxymethyl, 2-furylcarboxymethyl, 3-furylcarb-oxymethyl, a-sulfaminobenzyl, 3-thienylsulfaminomethyl, (x- (N-methylsulfamino) -benzyl, D (-) - 2-thienylguanidinome- 65thyl, D (-) - a-guanidinobenzyl, a-guanylureidobenzyl, a-hy-droxybenzyl, a-azidobenzyl, a-flurobenzyl, 4- (5-methoxy-l, 3-oxadiazolyl) aminomethyl, 4- (5-methoxy-l, 3-oxadiazolyl) -

hydroxy methyl, 4- (5-methoxy-l, 3-sulfadiazolyl) hydroxymethyl, 4- (5-chlorothienyl) aminomethyl, 2- (5-chlorothienyl) hydroxymethyl, 2- (5-chlorothienyl) carboxymethyl , 3- (l, 2-Thi-azolyl) aminomethyl, 3- (l, 2-thiazolyl) hydroxymethyl, 3- (l, 2-thiazolyl) carboxymethyl, 2- (l, 4-thiazoIyl) aminomethyl , 2- (l, 4-thiazolyl) hydroxymethyl, 2- (l, 4-thiazolyl) carboxymethyl, 2-benzothienylaminomethyl, 2-benzothienylhydroxymethyl, 2-benzothienylcarboxymethyl, a-sulfobenzyl, a-phosphonobenzyl, a -Diethylphosphono and a-monoethyl-phosphono.

Other interesting acyl residues in this class where X = oxygen are:

O

II

-CCHR4R5,

wherein R4 and R5 are defined below. R4 means hydrogen, halogen, such as chlorine, fluorine, bromine, iodine, amino, guanidino, phosphono, hydroxy, tetrazolyl, carboxy, sulfo or sulfamino and R5 means phenyl, substituted phenyl, a mono- or bicyclic heterocyclyl containing one or more oxygen -, Sulfur or nitrogen atoms in the ring, such as furyl, quinoxalyl, thienyl, quinolyl, quinazolyl, thiazolyl, isothiazolyl, tetrazolyl, oxadiazolyl, thiadiazolyl and the like, substituted heterocycles, phenylthio, phenyloxy, lower alkyl with 1 to 6 carbon atoms or substituted heterocyclic heterocyclic thio groups or cyano. The substituents on the groups R4 and R5 can be halogen, carboxymethyl, guanidino, guanidinomethyl, carboxyamidomethyl, aminomethyl, nitro, methoxy or methyl. R4 is selected from the group consisting of hydrogen, hydroxy, amino or carboxy and R5 is selected from the group consisting of phenyl or a 5- or 6-membered heterocyclic ring with one or two sulfur, oxygen or nitrogen heteroatoms, such as tetrazolyl, thienyl, Furyl and phenyl, the following acyl radicals are typical examples: phenylacetyl, 3-bromophenylacetyl, p-aminomethylphenylacetyl, 4-carboxymethylphenylacetyl, 4-carboxyamidomethyl-phenylacetyl, 2-furylacetyl, 5-nitro-2-furylacetyl, 3-furylace- tyl, 2-thienylacetyl, 5-chloro-2-thienylacetyl, 5-methoxy-2-thienylacetyl, a-guanidino-2-thienylacetyl, 3-thienylacetyl,

2- (4-methylthienyl) acetyl, 3-isothiazolylacetyl, 4-methoxy-3-isothiazolylacetyl, 4-isothiazolylacetyl, 3-methyl-4-isothiazolyl-acetyl, 5-isothiazolylacetyl, 3-chloro-5-isothiazolylacetyl, 3- Methyl-1,2,5-oxadiazolylacetyl, 1,2,5-thiadiazolyl-4-acetyl,

3-methyl-l, 2,5-thiadiazolylacetyl, 3-chloro-l, 2,5-thiadiazolyl-acetyl, 3-methoxy-l, 2,5-thiadiazolylacetyl, phenylthioacetyl,

4-pyridylthioacetyl, cyanoacetyl, 1-tetrazolylacetyl, a-fluoro-phenylacetyl, D-phenylglycyl, 4-hydroxy-D-phenylglycyl, 2-thienylglycyl, 3-thienylglycyl, phenylmalonyl, 3-chlorophenylnylmalonyl, 2-thylmalonyl, 2-th Thienylmalonyl, a-phosphonophenylacetyl, a-aminocyclohexadienylacetyl, a-sulfamino-phenylacetyl, a-hydroxyphenylacetyl, a-tetrazolylphenyl-acetyl and a-sulfophenylacetyl.

The acyl radical can also be selected from sulfur (1) and phosphorus (2) radicals:

(O) m (X) n

II I

-S — Y ° -P-Y '

(O) n 1

Y »

2nd

wherein with respect to 1_, m and n are integers which are selected between 0 and 1 and Y ° = 0® M®, -N (R ") 2 and R"; where M® is selected from hydrogen, alkali cations and organic

633 800

see bases and R "has the meaning given above, for example alkyl, alkenyl, aryl and heteroaryl. With regard to 2, X = 0 or; n = 0 or 1; and Y 'and Y" are selected from the groups consisting of 0® M ®, -N (R ") 2, R" and ZR ", in which all symbols have the meaning given above, and for example R" and ZR "typically have the following meaning: alkyl, alkenyl, aryl, heteroaryloxy, Y 'and Y ", including the R" groups, which can be linked to form cyclic esters, ester amide and amide functions. Typical examples of 1 are 0- (methylsulfonyl) thienamycin, O- (p-nitrophenylsulfonyl) - thienamyein, 0- (p-chlorophenylsulfinyl) thienamycin, 0- (o-Ni-trophenylsulfenyl) thienamycin, O-sulfamoylthienamycin, O-dimethylsulfamoylthienamycin and the sodium salt of thienamycin-O-sulfonic acid. Typical examples of 2 are 0- (Di -methoxyphosphino) -thienamycin, O-dibenzyloxyphosphino) -thienamyein, 0- (dihydroxyphosphino) -thienamycin disodium salt, 0- ( Dimethoxyphosphinyl) thienamycin, 0- (dimethoxyphosphinothioyl) thienamycin, O- (dibenzyloxyphosphinyl) thienamyeine and 0- (dihydroxyphosphinyl) thienamycin indium trium salt. The definition of Rr and R2 'does not include residues 1 ^ and 2.

An acyl class of particular interest are those acyl radicals which are selected from the group consisting of customary known N-acyl blocking or protecting groups, such as carbobenzyloxy, ring-substituted carbobenzyloxy, such as o- and p-nitrocarbobenzyloxy, p-methoxycarbobenzyloxy, chloroacetyl , Bromoacetyl, phenylacetyl, tert-butoxycarbonyl, trifluoro-acetyl, bromoethoxycarbonyl, 9-fluorenylmethoxycarbonyl, di-chloroacetyl, o-nitrophenylsulfenyl, 2,2,2-trichloroethoxycarb-onyl, bromotert-butoxycarbonyl, phenoxyacetyl; Non-acylated protective groups such as tri-lower alkylsilyl, for example tri-methylsilyl and tert-butyldimethylsilyl are also of interest.

The following radicals are, in accordance with the abovementioned definition of acyl, particularly preferred for R3 of structure IIa: formyl, acetyl, propionyl, butyryl, chloroacetyl, methoxyacetyl, aminoacetyl, methoxycarbonyl, ethoxycarb-onyl, methylcarbamoyl, ethylcarbamoyl, phenylthiocarbonyl, 3-aminopropion 4-aminobutyryl, N-methylaminoacetyl, N, N-dimethylaminoacetyl, N, N, N-trimethylaminoacetyl, 3- (N, N-dimethyl) aminopropionyl, 3- (N, N, N-trimethyl) aminopropionyl, N, N, N-triethylaminoacetyl, pyridiniumacetyl, guanidinoacetyl, 3-guanidinopropionyl, N3-methylguanidino-propionyl, hydroxyacetyl, 3-hydroxypropionyl, acryloyl, propynoyl, malonyl, phenoxycarbonyl, amidinoacetyl, acetylamido, acetylamide, acetamididinoid, acetamididinoid ureidoacetyl, guanylcarbamoyl, carboxymethylaminoacetyl, sulfoacetylaminoacetyl, phosphonoacetylaminoacetyl, N3-dimethylaminoacetamidinopropionyl, ureidocarbonyl, dimethylaminoguanylthioacetyl, 3- (l-methyl-4-pyrionimine) zol-1 -yl) -propionyl, 3-methyl-1-imidazolium acetyl, 3-sydnonylacetyl, o-aminomethylbenzoyl, o-ami-nobenzoyl, sulfo, phosphono,

Another class of acyl radicals are, for example, terminally substituted acyls in which the substituent is a basic group which can be substituted or unsubstituted: amino, amidino, guanidino, guanyl and nitrogen-containing 5 de mono- and bicyclic heterocycles (aromatic and non-aromatic) in which the heteroatom or the heteroatoms are selected from oxygen and sulfur in addition to the nitrogen. Such substituted acyls can be represented by the following formula:

10th

O

-C (CH2) m-A- (CH2) n -Y °,

characterized in which m and n are integers from 0 to 15 5;

A is 0, NR '(R' is hydrogen or lower alkyl of 1 to 6 carbon atoms), S or A is a single bond; and Y ° is selected from the following groups:

1.) Amino or substituted amino:

20th

—N (R °) 2 and -N (R °) 3,

wherein the values for R ° are independently selected from: hydrogen; N (R ') 2 (R' is hydrogen or lower alkyl of 1 to 6 25 carbon atoms); Lower alkyl and lower alkoxyl with 1 to 6 carbon atoms; Lower alkoxy lower alkyl, wherein the alkoxyl group comprises 1 to 6 carbon atoms and the alkyl group has 2 to 6 carbon atoms;

Cycloalkyl and cycloalkylalkyl, wherein the cycloalkyl group 30 comprises 3 to 6 carbon atoms and the alkyl group has 1 to 3 carbon atoms, and where two R ° groups can be connected to one another via the N atom to which they are bonded to form a ring with 3 to 6 atoms.

35

2.) Amidino and substituted amidino:

—N — C — N (R °) i,

I.

R °

wherein the values for R ° are independently selected from the groups: hydrogen; N (R ') 2 (R' is hydrogen or lower alkyl of 1 to 6 carbon atoms); Lower alkyl and lower alkoxyl of 1 to 6 carbon atoms, lower alko-45 xy lower alkyl, wherein the alkoxyl group contains 1 to 6 carbon atoms and the alkyl group contains 2 to 6 carbon atoms (if the lower alkoxy lower alkyl group is attached to a carbon atom, the alkyl group comprises 1 to 6 carbon atoms); Cycloalkyl and cycloalkylalkyl, wherein the alkyl group 50 comprises 1 to 3 carbon atoms; two R ° groups can be connected to each other with the atom to which they are attached to form a ring with 3 to 6 atoms;

3.) Guanidino and substituted guanidino:

55

O

II

P (OCH3) 2,

S

II

-P [N (CH3) 2] 2,

S

II

-P-N (CH3) 2 XDNa

-P (OCH3) 2,

O

-R

/ OCH3

-P-N (CH3) 2 ONa

ONa

O

II

-P [N (CH3) 2] 2,

-NH-C-N (R °) 2, I

NO

where R ° has the definition given under 2. 60 4. Guanyl and substituted guanyl:

~ C = NR,

I.

N (R °) 2

where R ° has the definition as previously given in 2.

5.) Nitrogen-containing mono- and bicyclic heterocycles (aromatic and non-aromatic) with 4 to 10 kernato

65

9

633 800

men, in which the heteroatom or the heteroatoms other than nitrogen are selected from oxygen and sulfur. Such heterocycles are typically illustrated by the following list of residues (R 'is H or lower alkyl with 1 to 6 carbon atoms):

Cycloalkyl and cycloalkylalkyl, wherein the cycloalkyl group comprises 3 to 6 carbon atoms and the alkyl group has 1 to 3 carbon atoms, and where two R ° groups can be connected to one another via the N atom to which they are bonded to form a ring with 3 to 6 atoms.

2.) Amidino and substituted amidino:

—N = C-N (R °) 2,

i

R °

wherein the values for R ° are independently selected from the groups: hydrogen; N (R ') 2 (R' is hydrogen or lower alkyl of 1 to 6 carbon atoms); Lower alkyl and lower alkoxyl having 1 to 6 carbon atoms, lower alkoxy lower alkyl, wherein the alkoxyl group contains 1 to 6 carbon atoms and the alkyl group contains 2 to 6 carbon atoms (if the lower alkoxy lower alkyl group is attached to a carbon atom, the alkyl group comprises 1 to 6 carbon atoms); Cycloalkyl and cycloalkylalkyl, wherein the alkyl group comprises 1 to 3 carbon atoms; two R ° groups can be connected to each other with the atom to which they are attached to form a ring with 3 to 6 atoms;

3.) Guanidino and substituted guanidino:

-NH-C-N (R °) 2,

j, "<

-O

<y

+ •

25th

30th

NO

where R ° has the definition given under 2. 4.) Guanyl and substituted guanyl:

-C = NO

35

40

NO%

where R ° has the definition as previously stated in 2.

5.) Nitrogen-containing mono- and bicyclic heterocycles (aromatic and non-aromatic) with 4 to 10 core atoms, in which the hetero atom or the hetero atoms are selected from nitrogen and oxygen apart from nitrogen. Such heterocycles are typically illustrated by the following list of residues (R 'is H or lower alkyl with 1 to 6 carbon atoms):

50

-O

(r *)

The following specific acyl residues that fall into this class are also representative:

o nh

II I

-c ch2ch2nhc-ch3

o nh

II I

-cch2ch2nhc-h o nh

II I

-cch2ch2nhc-nh2

O

II

-cch2ch2n (ch3) 2

O

II e

-cch2ch2n (ch3) 3

o nh

II I

-cch2ch2c-nh2

o nh

II I

-c ch2ch2ch2nhc-ch3

O

II

-cch2ch2ch2n (ch3) 3

O

II

-cch2ch2ch2n (ch3) 2

o vo

^ s-r.

* r \

J

60

-CCTUS-C

O

^ NH

-1

XNH,

-cch2s-ch2-c;

^ NH

O

-cch, -0-ch, c

\ nh2

^ NH \ nh7

633 800

10th

Production of the starting materials Ia, Ib, Ic, Id and Ie

The starting materials described above can be easily prepared from an N-protected thienamycin (JJ, such as an N-acylated thienamycin (1),

Th-

OH

nrrr2 'COOH

wherein R1 'and R2' are selected from hydrogen and the aforementioned acyl radicals. Preferably Rr is hydrogen and R2 'is an easily removable blocking group such as: carbobenzyloxy, ring substituted carbobenzyloxy such as o- and p-nitrocarbobenzyloxy, p-methoxycarbobenzyloxy, chloro-acetyl, bromoacetyl, phenylacetyl, -t-butoxycarbonyl, trifluoro-acetyl , Bromoethoxycarbonyl, 9-fluorenylmethoxycarbonyl, dichloroacetyl, o-nitrophenylsulfenyl, 2,2,2-trichloroethoxycarbonyl, bromot-butoxycarbonyl, phenoxyacetyl; Non-acyl-protected groups, such as tri-lower alkylsilyl, for example tri-methylsilyl and tert-butyldimethylsilyl, are also of interest. The most preferred N-blocking groups are substituted and unsubstituted carbobenzyloxy:

O

R2 = -C — OCH2

<1

r '

(R ') ",

wherein n is 0 to 2 (n = 0, R '= hydrogen) and R' is lower alkoxy or nitro; and bromo-tert-butoxycarbonyl,

O

CH

r

-C-O-C — CH2Br \

ch3

The final N deblocking in the manufacture of Ia, Ic or Ie can be carried out by a variety of known methods, including hydrolysis or hydrogenation; when hydrogenation is used, suitable conditions include a solvent such as a lower alkanol in the presence of a hydrogenation catalyst such as palladium, platinum or oxides thereof.

5 The N-acylated intermediate [1, (or Ia)] is prepared, for example, by treating thienamycin (I) with an acylating agent, for example an acyl halide or an acyl anhydride, such as an aliphatic, aromatic, heterocyclic, araliphatic or heterocyclic io aliphatic carboxylic acid halide or anhydride. Other acylating agents which can also be used are, for example, mixed carboxylic anhydrides and in particular lower alkyl esters of mixed carboxylic carboxylic acid anhydrides; carboxylic acids in the presence of a carbodi-15 mide such as 1,3-dicyclohexylcarbodiimide and an activated ester of carboxylic acids such as p-nitrophenyl ester are also suitable.

These N-acylated thienamycin starting materials are fully described, for example, in U.S. Patent Application No. 20 Serial No. 634,291 dated November 21, 1975 and the Continuation-in-part U.S. Application, which coexists with the present application for the same Applicant has been filed with the internal designation 15775YA. These applications are hereby incorporated into the description.

The acylation reaction can be carried out at temperatures in the range from about -20 to about 100 ° C, but preferably temperatures in the range from -9 ° C to 25 ° C are used. Any solvent in which the reactants 30 are soluble and which is substantially inert can be used, for example polar solvents such as water, alcohols and polar organic solvents, generally such as dimethylformamide (DMF), hexamethylphosphoramide (HMPA), acetone , Dioxane, tetrahydrofuran (THF), Acetoni-35 tril, heterocyclic amines, such as pyridine, ethyl acetate, aqueous mixtures of the above, and also halogenated solvents, such as methylene chloride and chloroform. The reaction is carried out for a time ranging from about 5 minutes to a maximum of 3 hours, but generally a reaction time of about 0.5 to about 1 hour is sufficient. The following equation can describe this method using a carboxylic acid halide; however, it goes without saying that the same products can be obtained by using a carboxylic acid anhydride or another functional equivalent acylating agent:

Oh oh

-N_ I

.sch, ch2-nh2 cooh acyl halide

_x \

i! |

-n L-cooh sch2ch2nr1ir2 '

In general, if an acid halide (suitable halides are chlorides, iodides or bromides) or an acid anhydride is used in the acylation reaction described above, the reaction is carried out in water or in an aqueous mixture of a polar organic solvent, such as acetone, dioxane, THF, DMF, acetonitrile and the like in the presence of a suitable basic acceptor such as NaHC03, MgO, NaOH, K2HP04 and the like.

When carrying out the reaction described here, it is generally not necessary to protect the 2-carboxy group or the 1 'hydroxy group; in cases where the acylating agent is extremely water sensitive, it is sometimes advantageous to perform the acylation in a non-aqueous

55 solvent system. Triorganosilyl (or tin) derivatives of thienamycin quickly give the tris-triorganosilyl derivatives, for example tris-trimethylsilylthienamycin Th (TMS) 3:

Th-

OTMS NHTMS L COOTMS

65

These derivatives, which are readily soluble in organic solvents, are easily prepared by, for example, thienamycin with an excess of hexamate.

11

633 800

thyldisilazane and a stoichiometric amount of trimethylchlorosilane at 25 ° C with vigorous stirring under an N2 atmosphere. The resulting NH4C1 is removed, for example, by centrifugation and the solvent is removed by evaporation, giving the desired silyl derivative.

The intermediate starting product Ic is generally produced according to the following scheme; It should be noted, however, that direct esterification without protection of the amino group is also possible.

OH Th-4 NR'R2 'COOH

1

OH Th — 1- NR 'R2' COX'R3 '

Id

Deblocking

Th-

OH NH2 L COX'R3 'Ic

In the diagram above, all symbols generally have the meaning given above.

In general, the transition (L—> Ic) is effected according to customary prior art methods. These procedures include, for example:

1.) Reacting .1 (or I) with a diazoalkane such as diazomethane, phenyldiazomethane, diphenyldiazomethane, and the like in a solvent such as dioxane, ethyl acetate, acetonitrile and the like, at temperatures between 0 ° C and reflux for a period of a few Minutes to 2 hours.

2.) Reaction of an alkali salt of 1_ with an activated alkyl halide, such as methyl iodide, benzyl bromide or m-phenoxybenzyl bromide, p-tert-butylbenzyl bromide, pivaloyloxy methyl chloride and the like. Suitable reaction conditions include solvents such as hexamethylphosphoramide and the like, and temperatures from 0 ° C to 60 ° C for a period from a few minutes to 4 hours.

3.) Reaction of 1 ^ with an alcohol such as methanol, ethanol, benzyl alcohol and the like. This reaction can be carried out in the presence of a carbodiimide condensing agent such as dicyclohexylcarbodiimide and the like

30th

same. Suitable solvents at temperatures between 0 ° C and the reflux temperature and reaction times of 15 minutes to 18 hours include CHC13, CH3C1, CH2C12 and the like.

4.) Reaction of an N-acylated acid anhydride from JL, prepared by reacting the free acid 1 with an acid chloride such as ethyl chloroformate, benzyl chloroformate and the like, with an alcohol such as those listed under 3.) under the same reaction conditions as under 3 .) called. The anhydride is prepared by reacting 1_ and the acid chloride in a solvent such as tetrahydrofuran (THF), CH2C12 and the like at temperatures from 25 ° C to the reflux temperature for 15 minutes to 10 hours.

5.) Reaction of labile esters of such as trimethylsilyl ester, dimethyl tert-butylsilyl ester or the like with R3'X °, where X ° is halogen, such as bromine or chlorine, and R3 'has the meaning given above, in a solvent such as THF, CH2C12 and the like at temperatures between 0 ° C and reflux for 15 minutes to 16 hours. This implementation is carried out, for example, according to the following scheme:

Th-

OTMS

r OTMS

N-Acylation NHTMS 1 ^ Th-4-NR1 TMS

COOTMS

3rd

Esterification

COOTMS

TH-

OTMS

mild hydrolysis NR1 TMS —► TH-

COOR3 '

OH NHR1 L COOR3 '

wherein TMS means triorganosilyl, such as trimethylsilyl, and all other symbols have the meaning given above.

The amides of the present invention are usually prepared in a simple manner by reacting the acid anhydride (Ic, X '= O, R3' = acyl) with ammonia or with the selected amine, for example the aforementioned alkyl, dialkyl, aralkyl or heterocyclic Amines.

The previously outlined schemes for the esterification are known in the related bicyclic lactam tibiotics and in general in organic syntheses and are intended

OH

Th-4 NR'R2-

COXR Id

50

55

It is stated here "that, as a rule, no particular critical conditions are required with regard to the reaction parameters in the preparation of the N-acylated and carboxyl derivatives Ic which are suitable as starting compounds for the present invention.

The starting compounds Ia and Ie are generally prepared in a simple manner by any of the known esterification or etherification reactions of the secondary alcohol group of Id. Such processes include, for example:

OR3

> Th—- NR'R2

65

COXR Ie

1.) For the preparation of the ether embodiments, the acid catalyzed reaction of Id is usually carried out with an inert diazoalkane such as diazomethane, phenyldiazomethane, diphenyldiazomethane and the like

633 800

12th

Solvents such as dioxane, tetrahydrofuran (THF), halogenated hydrocarbons such as CH2C12, ethyl acetate and the like in the presence of a catalytic amount of a strong acid or Lewis acid such as toluenesulfonic acid, trifluoroacetic acid, fluoroboric acid, boron trifluoride and the like at temperatures between -78 ° C and 25 ° C for a few minutes to 2 hours.

2.) For the preparation of the ether embodiments, the reaction of Id with an alkylating agent, such as an active halide, for example methyl iodide, benzyl bromide, m-phenoxybenzyl bromide and the like, is usually carried out; Alkyl sulfonate such as dimethyl sulfate, diethyl sulfate, methyl fluorosulfonate and the like in the presence of a strong base capable of forming an alcoholate anion of Ib. Suitable bases include alkali and alkaline earth oxides and hydroxides, alkali alkoxides such as potassium tert-butoxide, tert. Amines such as triethylamine, alkali alkyls and aryls such as phenyllithium and alkali amides such as sodium amide. Suitable solvents include an inert anhydrous solvent such as tert-butanol, dimethylformamide (DMF), THF, hexamethylphosphoramide (HMPA), dioxane and the like, at temperatures from -78 ° C to 25 ° C for a period of a few minutes to 4 hours.

3.) For the preparation of the ester embodiments, the reaction of Id with any of the acyl radicals listed above usually takes place in its acid form. This reaction can be carried out in the presence of a carbodiimide condensing agent such as a dicyclohexylcarbodiimide and the like. Suitable solvents include an inert solvent such as CHC13, CH2C12, DMF, HMPA, acetone, dioxane and the like at temperatures from 0 ° C to

60 ° C for a period of 15 minutes to 12 hours.

Th-

or3

NRrR2 '

• Th-

COX'R3 Ie

OR3

nh2

COX'R3 'Ie

Th-

'or3

nh2

cooh Ia

10th

The starting material Ia is obtained simply and preferably when X 'is oxygen and R3' is an easily removable carboxyl protecting or blocking group (see above). The starting material Ia is produced, for example, by deblocking by a variety of known procedures including hydrolysis and hydrogenation. If the preferred carboxyl blocking groups are used (below), the preferred deblocking method is hydrogenation, in which the intermediate (Ie) is hydrogenated in a solvent such as a lower alkanol in the presence of a hydrogenation catalyst such as palladium, platinum or oxides thereof.

In this connection it should be noted that the suitable "blocking groups R3 'generally include the subgroups which are defined as previously as aralkyl, halo-alkyl, alkanoyloxyalkyl, alkoxylalkyl, alkenyl, substituted alkyl or aralkoxyalkyl and also alkylsilyl, in which the 25 alkyl group Has 1 to 10 carbon atoms. Suitable “blocking groups” R3 'include, for example, benzyl, phenacyl, p-nitrobenzyl, methoxymethyl, trichloroethyl, trimethylsilyl, tributyltin, p-methoxybenzyl and benzhydryl. These blocking groups are preferred because they are generally recognized as easily removable blocking groups in the prior art relating to the cephalosporins and penicillins.

The preferred carboxyl blocking groups are benzyl and substituted benzyl:

4.) For the preparation of the ester embodiments, Id is normally reacted with an acyl halide or an acid anhydride, the acyl groups having been previously described. In general, an acid halide is used in the aforementioned acylation reaction (suitable halides are chlorides, iodides or bromides or acid anhydrides) 40 and the reaction is carried out in an anhydrous organic solvent such as acetone, dioxane, methylene chloride, chloroform, DMF or the like in the presence of a suitable basic acceptors, such as NaHC03, MgO, triethylamine, pyridine and the like, at temperatures between 0 ° C to 40 ° C 45 for one hour to 4 hours.

Suitable acyl halides and anhydrides include: acetic anhydride, bromoacetic anhydride, propionic anhydride, benzoyl chloride, phenylacetyl chloride, azidoacetyl chloride, 2-thienylacetyl chloride, 2-, 3- and 4-nicotinyl chloride, p-nitro-50 benzoyl chloride, 2,6-dimethoxybenzoyl -Guanidino-phenylacetylchloride, methanesulfonylchloride, dibenzylphosphorchlorate, dimethylthiophosphorochloridate, 2-furoylethylcarboxylic anhydride, methylchloroformate, bis (p-nitrobenzyl) phosphorochloridate and the like.

r3 '= _ ch2-

V

(R ') n

55

5.) For the preparation of the ester embodiments, Id is usually reacted with a suitable substituted ketene or isocyanate, such as ketene, dimethyl ketene, methyl isocyanate, methyl isothiocyanate, chlorosulfonyl isocyanate and the like. Suitable solvents include dioxane, tetrahydrofuran, chloroform and the like, at temperatures from -70 ° C to 60 ° C for a period of 15 minutes to 18 hours.

Intermediate Ie is then normally N-de-blocked, as previously described, to give starting compounds Ie (Rr and R2 '= H) and Ia. Ia is normally made from Ie by deblocking the carboxyl group:

wherein n = 0 to 2 (n = 0, R '= H) and R' mean lower alkoxyl or nitro.

In an alternative procedure, the compounds of the present invention IIa can also be obtained by working on the substituted N-methylene thienamycin derivatives II to obtain the derivatives by introducing the group R3 and / or-COX'R3 '. Such a procedure is generally carried out in exactly the same way as described above, with the exception that the compound II replaces the previously described starting materials, such as Ia, Ic and Ie, and of course there is also no need to carry out an N deblocking.

Manufacture of guanidines

In general, the guanidines can be prepared simply by combining thienamycin or an O- or carboxyl derivative thereof (Ia, Ic or Ie, where R, '= R2' = H) with (a.) An -OR "(e.g. O-alkyl, O-aryl) pseudourea or an S-alkyl or S-aryl pseudothiourea; by reacting a substituted pseudourea with ammonia or an amino compound, such as an alkyl, aralkyl or heteroaralkylamine.

Suitable solvents for such reactions include water and buffered aqueous polar organic solvent mixtures with a pH of 7 to 9 or anhydrous polar organic solvents, such as dimethylformamide or

13

633 800

Hexamethylphosphoramide at a temperature of 0 ° C to 40 ° C for 1 to 24 hours.

Suitable reagents (a.) And (b.) Include:

(a.) -OR pseudoureas and -SR pseudothioureas

O-methyl pseudourea, S-methyl pseudothiourea, S-methyl pseudothionite urea, 0-2,4-dichlorophenyl pseudo urea, S-p-nitrophenyl pseudothiourea, 0-N, N-tri-methyl pseudourea and the like.

(b.) Amino reagents

The reaction, which includes the reagents (a.), Can be represented by the following diagram:

sch2ch2nh2

cox'r3

12 1

r r n-c = n-r

, nr r sch -) ch7n = c,, x nr h cox'r3

wherein R3, X ', R3', R1 and R2 have the meaning given above; X "is O or S and R" is as indicated and preferably lower alkyl or aryl.

The reaction, which includes the reagents (b.), Can be represented by the following diagram:

Nitrogen atom to which they are attached) 4 to 10 atoms, one or more of which can be a further heteroatom from the group consisting of oxygen, sulfur or nitrogen; R1 and R2 are substituted or unsubstituted, namely: Cy-5 ano; Hydrogen; Carbamoyl; Carboxyl; Alkoxycarbonyl and alkyl of 1 to 10 carbon atoms; Alkenyl of 2 to 10 carbon atoms; Alkynyl with 2 to 10 carbon atoms; Cycloalkyl of 3 to 10 carbon atoms; Cycloalkylalkyl and cycloalkylalkenyl having 4 to 12 carbon atoms; Cycloalkylene-nyl, cycloalkenylalkenyl and cycloalkenylalkyl having 3 to 10.4 to 12 or 4 to 12 carbon atoms; Aryl of 6 to 10 carbon atoms, aralkyl, aralkenyl and aralkynyl of 7 to 16 carbon atoms; mono- and bicyclic heteroaryl and heteroaralkyl having 4 to 10 ring atoms, one or 15 of which is a heteroatom from the group consisting of oxygen, sulfur or nitrogen, and in which the alkyl group of the heteroaralkyl radical contains 1 to 6 carbon atoms; mono- and bicyclic heterocyclyl and heterocyclylalkyl having 4 to 10 ring atoms, one or more of which is a heteroatom from the 20 group oxygen, sulfur or nitrogen, and in which the alkyl group of the heterocyclylalkyl radical contains 1 to 6 carbon atoms;

and wherein the aforementioned substituent or substituents on R1, R2 or on the ring formed by the compound 25 of R1 and R2 are selected from the group consisting of: chlorine, bromine, iodine and fluorine; Azido; Alkyl of 1 to 4 carbon atoms; Thio; Sulpho; Phosphono; Cyanothio (-SCN); Nitro; Cyano; Amino; Hydrazino; mono-, di- and trialkyl-substituted amino and hydrazino, wherein the alkyl has 1 to 6 carbon atoms; Hydroxyl; Alkoxyl of 1 to 6 carbon atoms; Alkylthio of 1 to 6 carbon atoms; Carboxyl, oxo; Alkoxycarbonyl having 1 to 6 carbon atoms in the alkoxyl group; Acyloxy of 2 to 10 carbon atoms; Carbamoyl and mono- and dialkylcarbamoyl, in which the alkyl group 35 pe each has 1 to 4 carbon atoms, and their pharmaceutically acceptable salts, is characterized in that thienamycin (I)

O i2

sch0chnnh = c-nr ~ r - ^ I

ox 'rj hnr1r2

Oh

/ X

G) i 2

-sch2ch2nh = c-nr ~ r .eoo3

^ z ^ cooh

(I)

no r

45 with intermediate protection of the oxygen functions and / or the carboxyl function, with a compound of the formula

X "R °

where all symbols have the meaning given above.

The process according to the invention for the preparation of guanidines of the general formula (II)

oa

£ ch-.ch-> N = c-X 2 z,

.cooh y

(II)

Û

wherein Y is -NR'R2 and X is -NR'R2, where

R1 and R2 are independently selected from hydrogen, nitro, hydroxyl, alkoxyl with 1 to 6 carbon atoms, amino, mono-, di- and trialkylamino, in which the alkyl groups each contain 1 to 6 carbon atoms; R1 and <R2 can be linked together to form a substituted or unsubstituted mono- or dicyclic heteroaryl or heterocyclyl containing (together with the

50 R'R2N-C = N-R '

wherein X "O or S and -X" R ° is the leaving group and R1 and R2 are as defined above and R ° is any residue, treated and compounds obtained, if any, converted into their pharmaceutically acceptable salts.

The products (II and IIa) can form a variety of pharmacologically acceptable salts such as acid addition salts, for example with hydrochloric acid, hydrobromic acid, sulfuric acid, nitric acid, p-toluenesulfonic acid 60 and methanesulfonic acid. The salts of this invention are pharmacologically acceptable non-toxic derivatives that can be used as active ingredients in suitable pharmaceutical dosage forms. They can also be combined with other drugs to form compositions that have a wide range of activities.

The new compounds which can be produced according to the invention are new valuable antibiotics which are generally active against various gram-positive and gram-negative bacteria

633 800 14

and therefore find the skin of the nose and the neck and bronchial tissues in human and veterinary medicine and find them. The compounds which can be prepared according to the invention can also be prepared in a simple manner in the form of powdery or can therefore be prepared as antibacterial medicaments for the treatment-liquid sprays for inhalations or in infections caused by gram-positive pastilles or preparations for brushing the throat . For ve and gram-negative bacteria, for example the medicinal use on the eyes or ears can be wise against Staphylococcus aureus, Escherichia coli, Klebsiella the preparations as special capsules in liquid or semi-pneumoniae, Serratia, Salmonella typhosa, Pseudomonas and are in liquid form or they can be in the form of drops and der-Bacterium proteus. The bacterial same that can be produced according to the invention can be used. For topical applications, rizide can also be used as additives for futan hydrophobic or hydrophilic bases as ointments, creams, agents and for food preservation and as formulations, lotions, ointments or powders. Disinfectant. For example, in addition to a carrier material, the compositions according to the invention in concentrations in the range from 0.1 to 50% can also contain other constituents, such as stabilizers, binders, antioxidants, preservatives, and a million parts of the antibiotic per million parts of solution , det, in order to contain the growth of harmful bacteria on lubricants, suspending agents, viscosity agents or medical and dental equipment to destroy flavors and the like. In addition, other active crop applications, for example in the case of components based on water, can also be present or inhibit and also as bactericides in industrial applications in the compositions, in order to use a broader spectrum of antibiotics in the white water of paper mills to see that to cause activity.

Inhibit growth of harmful bacteria.

The products that can be produced according to the invention can be used alone. The compositions can be used for veterinary medicine or in combination with active ingredients, for example as preparations that can be inserted within the breast in a large number of pharmaceutical preparations. with either a quick or slow release effect

These antibiotics and their corresponding salts can be formulated in

Forms of capsules or used as tablets, as powders or as liquid solutions or as suspensions and elixirs 25 The dosages to be administered are usually dependent. They can be administered orally, intravenously or intramuscularly to a significant extent depending on the condition of the treated. Patient from and on the weight of the patient, the administration

The compositions are preferably prepared in a sol delivery route and the frequency of administration, form being preferred to be preferred in the gastrointestinal tract absorptive route for general infections. Tablets and capsules for oral administration - 30 ^ unc ^ ^ ie oral route for infections of the intestine. The chung can be in single dosage form and can generally contain a daily oral dosage of about 2 to usual excipients, such as binders, for example about 600 mg of active ingredient per kg of body weight of Pa syrup, gum arabic, gelatin, sorbose, tragacanth or po- took one or more administrations per day. A lyvinylpyrrolidone; Fillers, for example lactose, sugar, preferred daily dosage for adults is in the range of corn starch, calcium phosphate, sorbose or glycerin; Lubricant 35 of about 15 to 150 mg of active ingredient per kg of body agent, for example magnesium stearate, talc, polyethylene weight.

lenglycol, silicon dioxide; Disintegrants, for example potato starch or suitable humectants, such as sodium lye. The present compositions can be in various sulfate. The tablets can be coated in a known manner, to which individual dosage forms are administered, for example. Oral liquid preparations can be in the form of aqueous, in solid or in liquid orally absorbable dosage solutions or oily suspensions, solutions, emulsions, syrups. The compositions per single dose, whether they are present in pen, elixirs and the like or can be in the form of dry liquid or solid, should generally be prepared from 0.1% to 99% of active kenes product so that they then contain material with water, where the preferred range between approximately one or another suitable carrier material before it is 10 and 60%. The compositions contained in the turn can be turned on. Such liquid preparations, generally from about 15 mg to about 1500 mg of active ingredients, can include conventional additives, such as suspending agents; in general, however, it is preferred to use a dosage mode of sorbose, syrup, methyl cellulose, glucose / sugar amount in the range from about 100 to 1000 mg. In the case of rup, gelatin, hydroxyethyl cellulose, carboxymethyl cellulose, parenteral administration, the single dose is contained in the all-aluminum stearate gel or hydrogenated edible oils, in general the pure compound in a slightly acidified, for example, almond oil, fractionated coconut oil, oil ester, Pro so aqueous sterile solution or Contained in the form of a soluble powder, pylene glycol or ethyl alcohol; Preservative that is intended to dissolve.

tel, for example methyl or propyl p-hydroxybenzoate or

Sorbic acid. Suppositories usually contain the usual- The following examples describe the invention,

Chen suitable substances for this, for example cocoa butter without restricting them. In the examples, one or the other glycerine. 55 thienamycin nucleus (I, as previously indicated) by the following

Compositions for injections can be prepared in single-dose form in ampoules or in containers with added preservatives for multiple doses. The compositions can be in the form of suspensions, solutions or emulsions in oil or aqueous carrier materials and they can contain formulating agents such as suspending agents, stabilizing agents and / or dispersing agents. Alternatively, the active ingredients can be in the form of powder so that they can be mixed with a suitable carrier material, for example sterile, aseptic water, before the 6S on which the secondary alcohol group, the amino group and application can be mixed. the carboxyl group are included. The inventive

The compositions can therefore also be described in suitable formable compounds in a simple manner as prepared for absorption by the slime as follows:

OH

Th-4 NH2 COOH

15

633 800

■ OH

Th-N = C-X

l-cooh

II

and

Th-

- OR3

-NH = C-X

Y

.COX'R3

Ila wherein X, Y, R3, X ', R3' and A have the meaning given above.

example 1

Production of N-guanylthienamycin

Thienamycin (11 mg) is dissolved in a pH 70 in phosphate buffer (1 ml) and adjusted to pH 8.3 with 0.1 in sodium hydroxide using an automatic delivery burette. To 5 of the magnetically stirred solution, 0-2,4,5-trichlorophenylisourea.hydrochloride (76 mg) is added in individual portions, the pH being kept almost constant by means of the automatic output burette. The reaction is carried out at 22 ° C. for 4 hours and the pH is then adjusted to 7.0 again with dilute acid. A sample of this solution, containing thienamycin and N-guanylthienamycin, is treated electrophoretically (50 V / cm, 25 minutes, pH 70, in phosphate buffer) and shows a positive Sakaguchi spray zone at 2.0 cm with respect to an anode and a positive one Ninhydrin spray zone 15 at 1.5 cm in the same direction.

Th-

OTMS

NHTMS or TH (TMS) 3 L COOTMS

TMS = TrimethyIsilyl

Example 4

C \

,. H-.NCN

Cl - <OV-CH 2

C-C

.HCl

Production of silyated thienamycin

Thienamycin (80.0 mg) is suspended in 40 ml of tetrahydrofuran (THF) under a nitrogen atmosphere and concentrated to 10 ml; add hexamethyldisilazane (1.0 ml) and trimethylchlorosilane (300 µl). The mixture is reacted for 20 minutes at 25 ° C. with vigorous stirring. The suspension is then centrifuged to remove ammonium chloride. The supernatant is evaporated to an oil for further reaction under a stream of nitrogen.

Example 2

OH

Th-- NHC

^ NH

\

NH,

l co2h

35

40

45

Production of N-guanylthienamycin

Thienamycin (8.9 mg) is dissolved in pH 70 in phosphate buffer (0.7 ml) and N, N-dimethylformamide (0.3 ml) and the solution is brought to 50 pH 9 with the addition of 2.5N sodium hydroxide solution. 5 brought. O-methylisourea.hydrogen sulfate (43 mg) is added to the magnetically stirred solution, causing a slight drop in pH. Additional sodium hydroxide solution is added, adjusting the pH back to 9.5 and the solution is stirred at 23 ° C for 30 minutes. 55 The solution is then acidified to pH 7.0. A sample of the solution containing a mixture of thienamycin and N-guanylthienamycin shows two bioactive zones after electrophoresis (50 V / cm, 20 minutes, 0.05n pH 7 phosphate buffer) and bioautography against S. aureus plates. 60

Example 3

CÏ

Thienamycin C1_ ^ q \ _q_c ^

• HCl

Cl

VNH_

Th-

Oh

NHC ^ CO, H

^ NH

65

NH,

Production of 0-2,4,5-trichlorophenylisourea.Hydrochloride

A solution of cyanamide (0.28 mg) in ether (0.50 ml) is mixed with 2,4,5-trichlorophenol (12.5 g). The mixture is heated to 70 ° C. and the melt is stirred magnetically while the reaction vessel is flushed with nitrogen. Then dry hydrogen chloride gas is slowly bubbled through the melt and the mixture is cooled to 22 ° C. The solid obtained is washed thoroughly with ether and filtered, giving 0-2,4,5-trichlorophenylisourea hydrochloride as a white solid F. 205 to 206 ° C.

Example 5 Preparation of N-Allylformamide

A mixture of allylamine (5.00 g, 87.6 mmol) and methyl format (5.26 g, 87.6 mmol) is stirred at 25 ° C. for 2 hours. A short distillation head is then added to the flask and the desired N-allylformamide is obtained as a colorless oil at 89 to 90 ° C (0.7 mm). Yield 7.0 g (94%). IR (CHC13) 3380.1680 cm "*; KMR (CHC13) Ö8.1 (1H, br s), 06.4-7.9 (1H, very wide), 05.5-6.3 (1H, m ), 04.9-5.5 (2H, m), 63.85 (2H, m).

Example 6

Production of ethyl allylimidate hydrochloride

Ethyl chloroformate (2.66 g, 24.47 mmol) are syringeed into N-allylformamide (2.08 g, 24.47 mmol) in a dry flask under N2. The mixture obtained is stirred for 2 hours at 25 ° C and during this time CO 2 develops very quickly. The reaction mixture is then heated to 45 ° C. until no further gas evolution occurs (2 hours). The viscous product is then cooled and treated in vacuo at 0.2 mm for 2 hours to remove all volatiles.

Example 7

Preparation of N- (l-buten-3-yl) formamide

A solution of 3.5 g (0.05 mol) of 3-amino-1-butene in 12 ml of methyl format is kept at 25 ° C. for 20 hours; the solution is then concentrated under reduced pressure to remove excess methyl format. The N- (1-buten-3-yl) formamide remaining as a residue is distilled under reduced pressure. A fraction is obtained which is 3 g

633 800

16

Contains N- (l-buten-3-yl) formamide (boiling point 58 to 60 ° C / 0.5 mm).

Example 8

Preparation of ethyl N- (l-buten-3-yl) formimidate

A mixture of 1.0 g of N- (l-buten-3-yl) formamide and one equivalent of ethyl chloroformate is stirred under N2 for 4 hours while CO 2 evolves. The mixture is stirred under reduced pressure for 3 hours to remove all unreacted ethyl chloroformate and the residue obtained is ethyl N- (l-buten-3-yl) formimidate.

Example 9

Preparation of methyl-N-dimethylaminoformimidate

To a stirred solution of N, N-dimethylform hydrazide (0.22 g) in 2.0 ml of chloroform is added methyl chloroformate (0.5 ml) under nitrogen. The mixture is heated to 40 ° C for 3 hours and then evaporated under nitrogen. The mixture is triturated with anhydrous ether. The supernatant solution is decanted and the residue is dried in a stream of nitrogen. Yield 284 mg. KMR CDC13ô, 9.13 (HC); 3.80 (OCH3), 3.01 (N (CH3) 2).

Example 10

Manufacture of cyclopropylformamide

A mixture of cyclopropylamine (5.00 g, 87.6 mmol) and methyl format (5.26 g, 87.6 mmol) is stirred at 25 ° C. for 2 hours. An exothermic reaction is initially observed. The mixture is then placed on a rotary evaporator to remove the methanol formed during the reaction. The remaining material is distilled through a short distillation head to give 6.92 g (93%) of the desired N-cyclopropylformamide as a colorless oil. KMR (CDCl3) ô8, l (IH, br S); 06.8-8.5 (IH, br), 02.4-3.0 (IH, m), 00.4-1.0 (4H, m).

Example 11

Production of ethyl-N-cyclopropylformimidate

Ethyl chloroformate (4.078 g, 37.58 mmol) is syringed into N-cyclopropylformamide (3.194 g, 37.58 mmol) under nitrogen in a dry flask. Rapid gas evolution begins after an initial time of 30 seconds. The reaction mixture obtained is stirred at 25 ° C. until no further gas evolution can be detected (~ 4 hours) and then the viscous product is exposed to a vacuum of 0.5 mm for 1 hour to remove unreacted ethyl chloroformate. KMR analysis of the product shows the formyl proton at 69.37 as a broad singlet (CDC13 solution).

Example 12

Preparation of ethyl N- (methylthioethyl) formimidate

To a 60 ml separatory funnel are added ethyl formimidate (0.97 g, 8.8 mmol), a solution of ß-methylthioethylamine (0.80 g, 8.8 mmol) in CH2C12 (35 ml) and H2Ö (35 ml). The mixture is shaken vigorously for 5 minutes. The CH2C12 layer is separated, washed with brine and dried over magnesium sulfate, filtered and evaporated under reduced pressure to give the crude imidate (0.59 g) as a cloudy, pale yellow liquid. IR 1660, 1230 cm-1.

Example 13 Preparation of N-nitroguanylthienamycin

Thienamycin (131 mg) is in a solution of dimethyl

sulfoxide (10 ml), tri-n-butylamine (0.30 ml) and 2-methyl-1-nitro-2-thiopseudourea (0.3 g) dissolved. The solution is warmed to 45 ° C on a water bath while vigorously bubbling a stream of nitrogen through the solution. After 50 minutes, the solution is concentrated to 1.0 ml under high vacuum and then dissolved in 0.05N pH 7 phosphate buffer (7 ml). The unreacted thiopseudourea is precipitated and removed by filtration. The solution is then chromatographed over Dowex 50-Xa resin (53 cm3,200-400 mesh, Na + cycle) and eluted with water. The N-nitroguanyl derivative elutes in the first column volume and is freeze-dried to a solid (23%).

UV (pH 7 0, in phosphate buffer) ^ max 269 nm (express 000) electrophoresis (40 V / cm, pH 7 0, in phosphate buffer, 20 15 minutes) 3.0 cm in the direction of the cathode.

Example 14

Production of ethyl-N-methoxyformimidate

A mixture of methoxyamine hydrochloride (0.020 mol, 201.6700 g) and anhydrous potassium carbonate (0.01 mmol), 1.3821 g) is dissolved in 7.0 ml of water. 80 ml of ether are added and the reaction mixture is treated with ethyl formimide hydrochloride (0.02 mol, 2.1900 g). The mixture is shaken for 15 minutes. The ether layer is separated and the aqueous layer is extracted with 2 parts of ether (30 ml). The combined and dried ether solutions are evaporated to give 0.8433 g of ethyl N-methoxy formimidate.

KMR 61.36 (triplet)

30 63.83 (singlet)

64.13 (quartet)

66.56 (singlet).

Example 15

35 ethyl N- (2,2,2-trifluoroethyl) formimidate

Ethyl formimidate hydrochloride (0.555 g, 5 mmol), 2,2,2-trifluoroethylamine hydrochloride (0.677 g, 5 mmol) and potassium carbonate (0.345 g, 2.5 mmol) are suspended in 20 ml CH3Cl2 and treated with 2 ml H20. The mixture is shaken vigorously for 3 minutes. The organic phase is separated off and the aqueous phase is extracted twice with 10 ml portions of CH 2 Cl 2. The combined organic phases are dried and the CH2C12 is distilled off through a Vigreauxe column, giving ethyl N- (2,2,2-trifluoroethyl) formimidate. «KMR 61.33 t (CH3CH2); 3.8 q 0 = 10 c.p.s., CF3CH2) 4.23 q (u = 7.5, CH3CH2Ö); 7.6 S (H-C = N).

Example 16

Preparation of ethyl-N-ethoxycarbonylethyl formimidate 50 ethyl formimidate hydrochloride (0.55 g, 5 mmol), ethyl glycine nathydrochloride (0.697 g, 5 mmol) and potassium carbonate (0.345 g, 2.5 mmol) are suspended in 20 ml of CH2C12 and with 2 ml H20 treated. The mixture is shaken vigorously for 4 minutes. The organic phase is separated, the aqueous phase extracted twice with CH2C12 (10 ml) and the combined organic phase is dried and evaporated to give ethyl N-ethoxycarbonylmethylformimidate.

KMRÔ: 1.26 t (CH2-CH2); 4.06 S (N-CH2-C); 4.23 g (CH3CH2-Ö); 7.5 S (N = CH).

60

Example 17

Production of potassium N-ethoxycarbonylmethylformimidate

H

\

/

C = N-CH, • COOEt

KOEt,

Hv

EtO

Et = C2H5

Et2ö, EtOH

EtO

JC = N-CTL-COOK

17th

633 800

Potassium (0.18 g) is mixed with 0.6 g of ethanol and

4 ml of ethyl ether dissolved under N2. The solution is diluted with 50 ml of ethyl ether and ethyl N-ethoxycarbonylmethylformi-midate (0.79 g) in 2 ml of ethyl ether is added, followed by 0.1 ml of HzO. Rapid crystallization of the salt takes place. The solid is filtered, washed with ether and dried under vacuum to give potassium N-ethoxycarbonylmethyl formimidate.

KMR (D20) 1.13 t (CH3CH2); 3.63 g (CH3-CH20); 3.8 S (N-CH2-C); 8.06 S N = CH.

Example 18 Preparation of ethyl N-benzyl formimidate

A solution of 690 mg (5.1 mmol) of N-benzylformamide in

5 ml of methylene chloride is cooled on an ice-water bath and placed under argon protection. The solution is stirred while 4.9 ml (4.9 mmol) of 1M triethyloxonium fluoroborate in methylene chloride are added dropwise. After a reaction time of 45 minutes, the mixture is concentrated to dryness under reduced pressure at room temperature and the residue is dried under reduced pressure over P2Os. The nuclear magnetic resonance spectrum of the product in deuterochloroform is completely in agreement with the fluoroborate etherate complex of ethyl-N-benzylformimidate.

Example 19

Production of N-isopropylformamide

Formamide (1.13 g, 0.98 ml) is dissolved in 10 ml of toluene containing toluenesulfonic acid (4.7 g. To this mixture is added isopropylamine (2.95 g, 4.25 ml). The mixture is poured over The mixture is filtered and the toluene evaporated under reduced pressure and the residual oil is distilled at 59-62 ° C / 0.07 mm to give 1.0 g of the desired product.

Example 20

Preparation of methyl N-isopropyl formimidate

Isopropylformamide (535 mg) is treated with the equivalent amount of ethyl chloroformate (440 ul) for 2 to 3 hours at 40 to 45 ° C under nitrogen. The mixture is washed successively with petroleum ether and anhydrous ether and benzene to give the product as an oil.

Example 21

Following the procedure as described in the preceding text and in the examples, the following compounds of the invention can be obtained. The reagents, imido ethers and imidohalides used for the reaction with thienamycin or a derivative thereof for the preparation of the compounds listed below are either known or can be prepared in the manner described above.

OH

ch? ch 2n = c-n

/ R

<f

-COOH

R

V

Ver

binding r

r1

r2

170)

nh2

ch3

H

171)

nh2

ch3

ch3

172)

nhch3

ch3

ch,

173)

n (ch3) 2

ch3

ch3

174)

nh2

c2h5

H

175)

nh2

-ch (ch3) 2

H

176)

nh2

-chch = ch2

H

177)

178)

179)

180)

nhch3

nhch3

nh2

nh2

-ch (ch3) 2

ch3

n (ch3) 2

nhnh2

h h h

H

Manufacture of alternative raw materials

In addition to thienamycin, it is obvious to the person skilled in the art

that its various isomers, alone or as mixtures, can be used as starting materials in the preparation of the compounds which can be prepared according to the invention. Some of these isomers are available, for example, from natural products of fermentation (see below) in the form of a mixture of 4 diastereoisomers which have bactericidal activity and which can be separated according to customary technical methods. The 4 diastereoisomers (2 eis, 2 trans) can be separated chromatographically. The separation of a given d / 1 pair with optically active acids or bases is carried out according to customary methods. It should be noted that the absolute configuration 20 of the first identified starting material (I) is 5R 6S 8R.

Production of thienamycin by total synthesis oh

25th

sch7ch2nh2 cooh

30th

Stage A. Preparation of 4- (2-acetoxyvinyl) azetidin-2 on-

H2C = CH-CH = CHOC-CH3 + O = C = N-SO2CI

35

40

O

0

ch = ch0cch,

ê

-ts \

0

ch = ch0cch3

45

s02ci

/

• nh

A solution of 1.0 ml of distilled chlorosulfonyl isocyanate (1.65 g, 11.7 mmol) in 2.5 ml of anhydrous diethyl ether is cooled under N2 in a bath at -20 ° C.

A solution of 2.5 g of L-acetoxybutadiene (22 mmol) in 2.5 ml of anhydrous ether is similarly cooled under nitrogen in a -20 ° C bath.

The chlorosulfonyl isocyanate solution is added dropwise to the acetoxybutadiene solution by means of a polytetrafluoroethylene tube immersed in the CSI-55 solution and is pressurized with N2. The addition takes 10 minutes. Little or no color is discernible and the reaction is stirred at -20 ° C for 0.5 hours. The solution is clear and has a light yellow color.

60 A solution of 2 g sodium sulfite and 5 g K2HPÖ4 in 20 ml H2Ö is prepared during the aforementioned 0.5 hour reaction time and cooled on an ice bath; 20 ml of ether are added to the mixture with vigorous stirring on an ice bath. After the end of the 30-minute reaction time, the reaction mixture is transferred to the vigorously stirred hydrolysis mixture, again using N2 pressure and a polytetrafluoroethylene tube, which is kept in a bath of −20 ° C. in the reaction flask. The snow

633 800

18th

le, dropwise addition is complete after 5 minutes. The hydrolysis is allowed to continue for another 5 minutes. The hydrolysis mixture has a pH of 6-8, preferably pH 8.

The phases are separated, with a yellow-orange resin remaining in the aqueous phase. The ether phase is dried directly over MgS04. The aqueous resin phase is extracted three times with 50 ml portions of ether, each portion being added to the starting ether / MgSO4 mixture.

The dried extracts are filtered and concentrated to 5 ml under a stream of N2; part of the product is crystallized at this stage.

A column of 10 g of Baker's silica gel, packed in ether, is made and the ether concentrate is added to the top and allowed to run through. The residue in the flask is rinsed three times with 2 ml of ether, pipetted off each time and added to the column. Then it is eluted with ether.

The first 25 ml are mainly empty volumes. The next 5 10 ml fractions are collected, followed by 3 50 ml fractions and all are reduced in volume under a stream of N2. The product crystallizes from fractions 4 to 6 with traces in 3 and 7. Fractions 1 to 3 contain a yellowish, pungent-smelling material that becomes resinous when standing. Yield: 100 mg as a mixture of the ice and trans isomers.

hours of stirring, the solution is neutralized with glacial acetic acid. Removal of the methanol in vacuo gives the crude 4- (2-hydroxyethyl) -2-azetidinone as an oil. The product is purified chromatographically on silica gel by eluting with 10% methanol / CHCl3 to give 1.55 g of the alcohol: mp 50 ° C .;

IR (CHC13) [x 5.67; KMR (CDC13) x 3.20 (broad s, 1, NH), 6.24 and 6.28 (m at t, total 3, C-4H and CH, OH corresponding), 6.90 (broad s at ' A AB pattern further splits into four by C-4H and NH, total 2, OH and C-3H respectively, Jgera = 13.0Hz, JViC = 4.2 Hz, JNH = 1.6 Hz), 7.42 ('A AB pattern further splits into four by C-4H and NH, 1, C-3H,

gladly

= 13.0Hz, Jvic = 2.2 Hz, JNH = 1.1 Hz), 816 (m, 2,

CH2CH2OH).

s

Level D:

Preparation of 8-oxo-2,2-dimethyl-3-oxa-l-azabicyclo [4,2,0] octane

Step B. Preparation of 4- (2-acetoxyethyl) -2-acetidinone

S

cch.,

c /

-nh

0

/

jra

A solution of 4- (2-acetoxyvinyl) -2-azetidinone (10.0 g,

0.065 mol) in 200 ml of ethyl acetate, containing 100 mg of a 10% palladium / C catalyst, is hydrogenated in a Parr shaker at 25 ° C. under a pressure of 2.8 kg / cm 2 over the course of 15 minutes. The mixture is filtered through a bed of Supercel and washed with additional ethyl acetate. The combined filtrate is washed in vacuo to give 4- (2-acetoxyethyl) -2-azetidinone (10.0 g) as a crystalline solid. When recrystallizing from ether, white crystals are obtained: mp 44 to 47 ° C;

IR (CHCl ^ 5.66.5.74; KMR (CDC13) x3.44 (broad s, 1, NH), 5.82 (m, 2, CH2OCOCH3), 6.29 (m, 1, C-4H , 6.87 ('A AB pattern is further divided into four by C-4H and NH, 1, Jgem = 12.8 Hz, J-4.5 HHnh = 1.9 Hz, 7.38 (' A AB pattern still divided into four by C-4H and NH,

1, Jgem = 12.8 Hz, J = 2.3 Hz, JNH = 1.0 Hz). 7.93 and 8.02 (s on m, total 5, OCOCHj and CH? CH7OCOCH3, respectively).

Step C. Preparation of 4- (2-hydroxyethyl) -2-azetidinone

A solution of 4- (2-hydroxyethyl) -2-azetidinone (1.87 g, 0.016 mol) and 2,2-dimethoxypropane (1.69 g, 0.016 mol) in 25 ml of anhydrous methylene chloride is quenched with boron trifluoride- ^ 30 etherate (0.201 ml, 0.002 mol) treated at 25 ° C. The obtained OCCH ^ ne solution is stirred for 10 minutes. When the solvent is removed under reduced pressure, an oil (2.5 g) is obtained. Chromatography of the crude product on a silica gel column using 2: 1 ethyl acetate / benzene 35 as eluent gives 8-oxo-2,2-dimethyl-3-oxa-l-azabi-cyclo [4,2,0] octane ( 1.59 g) as a crystalline solid. Recrystallization from ether / hexane gives a product with a melting point of 60 to 61 ° C.

IR (CHC13) ìx: 40 KMR (CDC13) t:

5.73 (ß-lactam)

6.02-6.28, m, 2H, C- ^ methylene

6.22-6.62, m, IH, C-6 methine

6.90, dd, 1H, J7j7 = 14 Hz, J6 7 = 4.5 Hz

C-7 proton ice to C-6H

7.47, dd, 1H, J7.7 = 14 Hz, J6.7 = 2 Hz

C-7 proton trans to D-6H

7.82-8.68, m, 2H, C-5 methylene

8.23, s, 3H 1. ,,. ,.

8.57, s, 3H) methyls

50

55

Level E: -

Preparation of 8-oxo-2,2-dimethyl-7a and ß- (l-hydroxy-ethyl) - (3-oxa-l-azabicyclo- [4,2,0] -octane oh

60 o

I.

"N.

S X

Under nitrogen at 0 ° C, a solution of 4- (2-aceto-xyethyl) -2-azetidinone (2.24 g, 0.014 mol) in 25 ml of anhydrous methanol with a solution of sodium methylate (77 mg, 1.4 mmol) ) treated in 5 ml of anhydrous methanol. After one-

A solution of 8-oxo-2,2-dimethyl-3-oxa-l-azabicyclo- [4, 2.0] octane in anhydrous tetrahydrofuran which has been cooled to -78 ° C was added. After 2 minutes, the lithium eno-

19th

633 800

lat treated with excess acetaldehyde. The solution is stirred at -78 ° C for 30 minutes and then poured into water. The aqueous phase is saturated with sodium chloride and extracted with ethyl acetate. The combined ethyl acetate solutions are dried over magnesium sulfate and filtered. The filtrate is evaporated under reduced pressure to give the crude product. Purification by means of chromatography on silica gel using ethyl acetate / benzene gives 8-oxo-2,2-dimethyl-7a and ß- (l-hydro-xyethyl) -3-oxa-l-azabicyclo- [4,2, 0] octane.

Data for 8-oxo-2,2-dimethyl-7ß- (l-hydroxyethyl) -3-oxa-l-azabicyclo [4,2,0] octane:

KMR (CDC13): 1.67, d, 2H, ArH 2.37, d, 2H, ArH 4.67, s, 2H, ArCH2 4.67-5.22, m, CH3CH 5 5.98-6, 25, m, 2H, C-4 methylene

6.25-6.62, m, IH, C-6 methine 7.75-8.83, m, 2H, C-5 methylene 8.22, s, 3H, C-2 methyl 8.50-8, 59, m, 5H, C-2 methyl + CH3CH

10th

The 7β-diastereoisomers or the la and 7ß-mixtures are obtained in an analogous manner.

IR (CH2C12) h: 5.12 \ x (β-lactam)

KMR 5.53-6.43, m, 4H, C-4 methylene +

(CDC13) t: C-6 methine + C-9 methine

6.90, dd on broad s, 2H, J7 9 = 9 Hz J6 7 = 5.5 Hz, C-7 methine + ÒH 7.70-8.83, m, 2H, C-5 methylene

8.27, s, 3HÌ £ _2 methyl 8.60, s, 3HJ ^ ^ lvlemyi 8.78, d, 3H, J9.10 = 6.5 Hz, C-10 methyl

Data for 8-oxo-2,2-dimethyl-7a- (l-hydroxyethyl) -3-oxa-1 -azabicyclo- [4,2,0] -octane:

IR (CHC13) [.i: 2.9 broad O-H

5.73 β-lactam KMR 4.23-3.33, m, C-6 methine

(Acetone - d6) x: 3.33, broad s, OH

2.83, dd, J = 2 Hz, 6 Hz \ _

2.67, dd, J = 2 Hz, 8 Hz J 1.93-1.63, m, C-5 methylene l, 40, 's} C ~ 2MethyIe i', 23, d, H = 6.5 Hz, C-10 methyl

Step F. Preparation of 8-oxo-2,2-dimethyl-7a- (l - ç> -ri \ trob <iïizy \ cs.Tb-onyldioxyäthyl) -3-oxa-l-azabicyclo-f4,2,0 ] octane

-7 methine

R * = ^ jOCH2-0

Level G:

15 Preparation of cis and trans-3- (l-p-nitrobenzylcarbonyldi-oxyethyl) -4- (2-hydroxyethyl) -2-azetidinone

OR

or oh

'X

.nh o

-R = -C-O-CH

1

/ îîo

"lïOr

Under anhydrous conditions, a solution of 8-oxo-2,2-dimethyl-7a (l-hydroxyethyl) -3-oxa-l-azabicyclo [4,2,0] octane (60 mg, 0.302 mmol) is dissolved in 0.6 ml ether treated with powdered potassium hydroxide (19 mg, 0.332 mmol). After 15 minutes, p-nitrobenzyl chloroformate (65 mg, 0.302 mmol) is added to the reaction mixture. The mixture is stirred at 25 ° C. for a further 15 hours. The mixture is divided between In pH 7 phosphate buffer and other ether. The ether phase is washed with water and brine, dried over magnesium sulfate and filtered. Evaporation of the filtrate under reduced pressure gives 67 mg of a colorless oil. Purification by preparative thick-layer chromatography over silica gel and development with 1: 9 ethyl acetate / benzene gives 8-oxo-2,2-dimethyl-7a- (lp-nitrobenzylcar-bonyldioxyethyl) -3-oxa-l-azabicyclo- [4,2,0] octane (40 mg) as a mixture of the diastereomers.

IR (CH2C12) [ì: 5.68 (β-lactam and carbonate), 6.19 and 6.54 (nitro)

8-Oxo-3-oxa-2,2-dimethyl-7a (lp-nitrobenzylcarbonyldi-oxyethyl) -l-azabicyclo- [4.2.0] octane (1.0 g) is dissolved in 8 ml of acetic acid and 2 ml of water dissolved and heated to 65 ° C for 30 minutes. The acetic acid and water are removed under reduced pressure and the residue is taken up in benzene and evaporated to give trans-3- (lp-nitrobenzylcarbonyldioxyethyl) -4- (2-hydroxyethyl) -2-azetidinone as a mixture of the diastereoisomers .

35 IR '(CH2C12) [ì: 5.67 (ß-lactam), 5.72 shoulder, 6.20 and 6.57 (nitro)

KMR (CDClj): 1.73, d, 2H, H = 8.5 Hz, ArH 2.43, d, 2H, J = 8.5 Hz, ArH "o 3.63, broad s, 1H, NH

4.37-5.13, m, 1H, CH3CH 4.72, s, 2H, ArCH2 6.07-6.53, m, 1H, C-4 methine 6.23, t, 2H, J = 5, 5 Hz, CH2OH 45 6.73-6.93, m, IH, C-3 methine

7.63-8.97, m, 3H, CH2CH2OH 8.53, d, J = 6.5 Hz, CH3CH The cis-diastereoisomers or the cis-trans mixture is obtained in an analogous manner.

Steps D ', E', F 'and G' as an alternative to steps D, E, F and G for the preparation of 3- (l-p-nitrobenzylcarbonyl-dioxyethyl) -4- (2-hydroxyethyl) azetidinone

50

55

60

OR

Oh

O'

-nh o ii

R = -COCH.

NO,

65

Levels D ', E, F and G'

Preparation of 1- (2-tetrahydropyranyl) -4- [2-tetrahydropyranyl) oxyethyl J-2-azetidinone

633 800

20th

A solution of 4- (2-hydroxyethyl) -2-azetidinone (62 mg, 0.539 mmol) in 0.5 ml of anhydrous p-dioxane with 2,3-dihydropyran (0.98 ml, 1.08 mmol) and p-toluosulfonic acid monohydrate (19 mg, 0.10 mmol) were treated. The resulting solution is stirred for 60 minutes and then partitioned between 10 ml of a 0.5m pH 7 phosphate buffer and 10 ml of ethyl acetate. The watery

Phase is extracted a second time with ethyl acetate. The combined ethyl acetate solutions are washed with salt water, dried over magnesium sulfate and filtered. The filtrate is evaporated under reduced pressure to give 216 mg of the crude product. Purification by preparative thick layer chromatography and development with ethyl acetate gives 80 mg of l- (2-tetrahydropyranyl) -4- [2- (2-tetrahydropyranyl) -oxyethyl] -2-azetidinone as an oil.

C-3 methylene io KMR 5.13-5.60, m, OCH (CDC13) t: 5.83-6.85, m, C- ^ H + OCH2 6.95, dd, J = 5 Hz and 15 Hz 7.35, dd, J = 3 Hz and 15 Hz 7.62-8.95, m, CHCH2CH2CH2CH2 + CHCH2CH20

15

Preparation of cis- and trans-1- (2-tetrahydropyranyl) -2- (1-hydroxyethyl) -4- [2- (2-tetrahydropyranyl) -oxyethyl] -2-azetidinone

If you work in the for the production of 8-oxo-2,2-dimethyl-7a and ß- (l-hydroxyethyl) -3-oxa-l-azabicyclo [4,2,0] octane from 8-oxo -2,2-dimethyl-3-oxa-l-azabicyclo- [4,2,0] -octane described manner and uses l- (2-tetra-hydropyranyl) -4- [2- (2-tetrahydropyranyl) -oxyethyl ] -2-acetidi-non, you get a diastereomeric mixture of both

Cis- and Trans-l- (2-tetrahydropyranyl) -3- (l-hydroxyethyl) -4- [2- (2-tetrahydropyranyl) oxyethyl] -2-azetidinone.

35 Preparation of cis and trans-l- (2-tetrahydropyranyl) -3- (l-p-nitrobenzylcarbonyldioxyäthyl) -4- [2- (2-tetrahydropyranyl) -oxyethylJ-2-azetidinone p *. = yes,

The procedure is the same as for the production of 8-oxo-2,2-dimethyl-7a- (lp-nitrobenzylcarbonyldioxyethyl) -8-oxa-l-azabicyclo [4,2,0] octane from 8-oxo -2,2 ethyl-7- (l-hydroxyethyl) -3-oxa-l-azabicyclo [4,2,0] octane and uses trans-l- (2-tetrahydropyranyl) -3- (l- hydroxyethyl) -4- [2- (2-tetrahydropyra-nyl) -oxyethyl] -2-azetidinone, there is obtained trans-l- (2-tetra-hydropyranyl) -3- (lp-nitrobenzylcarbonyldioxyethyl) -4- [2 - (2-tetrahydropyranyI) -oxyethyl] -2-azetidinone. The cis diastereoisomer is obtained in an analogous manner.

Preparation of cis and trans-3- (l-p-nitrobenzylcarbonyl-dioxyethyl) -4- (2-hydroxyethyl) -2-azetidinone or

h o-

.nh p. - c0ch., _ / y n0 "

21st

633 800

A solution of Trans-l- (2-tetrahydropyranyl) -3- (lp-nitrobenzylcarbonyldioxyäthyl) -4- [2- (2-tetrahydropyranyI) -oxyäthyI] -2-azetidinone in methanol at 25 ° C is with a 0 , 1 molar equivalent of p-toluenesulfonic acid monohydrate treated. The solution is stirred for 2 hours and then neutralized with pH 7 phosphate buffer. The product is extracted into ethyl acetate. The ethyl acetate solution is washed with salt water, dried over magnesium sulfate and filtered.

The filtrate is evaporated under reduced pressure to give trans-3- (l-p-nitrobenzylcarbonyldioxyäthyI) -4- (2-hydr-oxyäthyl) -2-azetidinone. The cis diastereoisomer is obtained in an analogous manner.

Level H:

Preparation of cis and trans-3- (l-p-nitrobenzylcarbonyldi-oxyethyl) -4- [2,2-bis- (2-hydroxyethyl] thioethyl] -2-azetidinone

/ "" "CHO

r '

V

25th

R = C0ZYL2- / y

> - »° 2

A mixture of anhydrous pyridine (0.146 ml, 1.81 mmol) and anhydrous, powdered chromium trioxide (92 mg, 0.916 mmol) in 8 ml of anhydrous acetonitrile is stirred under nitrogen at 25 ° C. for 30 minutes. 250 mg of dry supercel are added to the dark brown solution obtained, and then a solution of trans-3- (lp-nitro-benzylcarbonyldioxyethyl) -4- (2-hydroxyethyl) -2-azetidinone (186 mg, 0.550 mmol) in 1 ml anhydrous acetonitrile. After stirring for one hour, the reaction mixture is filtered through a mixed packed bed of 2 g each of silica gel and magnesium sulfate. The bed is washed repeatedly with a total of 30 ml of additional acetonitrile. The filtrate is concentrated under reduced pressure at 25 ° C to a volume of 3 ml. Thin layer chromatography (silica gel; ethyl acetate / benzene 2: 1) shows that this solution contains a product (Rf = 0.38) that is less polar than the starting material (Rf = 0.21).

The acetonitrile solution of trans-3- (lp-nitrobenzylcarbo-nyldioxyäthyl) -4- (2-oxoäthyl) -2-azetidinone, which was prepared as above, is under nitrogen at 0 ° C with 2-mercaptoethanol (0.386 ml , 5.5 mmol) and immediately thereafter treated with boron trifluoride etherate (0.176 ml, 1.43 mmol). After stirring for 15 minutes, this solution is divided between aqueous dipotassium hydrogen phosphate (1.5 g in 4 ml of water) and 12 ml of ethyl acetate. The aqueous phase is extracted a second time with ethyl acetate. The combined ethyl acetate solutions are washed with brine, dried over magnesium sulfate and filtered. The filtrate is evaporated under reduced pressure to give 229 mg of an oil. The

The product is purified by preparative thick layer chromatography on silica gel and developed with ethyl acetate, 30 mg of 118 mg of trans-3- (lp-nitrobenzylcarbonyldioxyethyl) -4- [2,2-bis (2-hydroxyethyl) thioethyl] -2-azetidinone as one colorless oil.

IR 5.75 (5.79 shoulder) β-lactam and carbonate

(CH2C12) | ì: 6.20.6.55 Nitro 3SKMR 1.70, d. J-8.5 Hz, 2H, ArH

(Acetone-d6) -c: 2.28, d, J = 8.5 Hz, 2H, ArH 2.48-2.88, m, 1H, NH 4.63, s, ArCH2

40

45

4.63-5.12, m, CH3CH

3H

5.80, t, J = 7 Hz, CH2CH

V

13H

50

5.80-7.45, m, C-4H + C-3H + SCH2CH2OH 7.63-8.33, m, 2H, CH2CH 8.53, d, J = 6.5 Hz, 3H, CH3CH

The cis diastereoisomers are prepared in an analogous manner. Alternatively, the mixed diastereoisomers are obtained if a mixture of the diastereoisomers is used as the starting material.

Level I:

55 Preparation of Trans-3- (l-p-nitrobenzylcarbonyldioxyethyl) -4- [2,2-bis- (2-azidoethyl) thioethyl] -2-azetidinone

OCOOPNB

, oh s ^ x /

. OH

sxx / + 2ch „s02c1 2et ^ ns nan ^ ^

0C00PN3

THF

N.

DMSO

f

_NH

633 800

22

To a solution of 211 mg (molecular weight = 474; 0.445 mmol) of trans-3- (lp-nitrobenzylcarbonyldioxyäthyl) -4- [2,2-bis- (2-hydroxyethyl) -thioethyl] -2-azetidinone in 5 ml of tetrahydrofuran ( THF) (distilled from lithium aluminum hydride) at 0 ° C, 103 mg of mesyl chloride (molecular weight = 114; 0.904 mmol) are added to 1 ml of tetrahydrofuran and immediately thereafter 134 ntriethylamine (molecular weight 101; e = 0.729; 0.967 mmol). The reaction mixture is stirred under N2 for 1 hour. The triethylamine hydrochloride is filtered under N2 and washed with a few ml of additional THF. The clear colorless filtrate is concentrated under a stream of N2 and then pumped under high vacuum for 10 minutes. The dimesylate is immediately dissolved in 5 ml DMSO (distilled from CaH2 at 8 mm and stored over a 4A Linde molecular sieve) in the presence of 347 mg NaN3 (molecular weight = 65; 5.34 mmol). After stirring overnight and under N2, 10 ml of H20 and 20 ml of ethyl acetate (EA) are added. The layers are separated and the aqueous layer is washed three times with 10 ml EA, each organic layer 5 being washed again with 10 ml H20 and 10 ml brine. The combined ethyl acetate layers are dried over anhydrous magnesium sulfate and filtered and concentrated under a stream of N2 to give the crude diazide. Preparative thin layer chromatography on Kie-îoselgel gives trans-3- (l-p-nitrobenzylcarbonyldioxyäthyl) -4- [2,2-bis- (2-azidoäthyl) -thioäthyl] -2-azetidinone. The cis-di-stereoisomers or the cis-trans mixture are obtained in an analogous manner.

is level J:

CO.K.

co2h c (ch) 2 -t 2n02_ ^ v \\ _ chn2 ea / etjo c0 „? nb

! z c (oh) 9 c02pnb

-f

0c00pnb

ch9ch., n,

^ J Toi. v sch7ch..nq: f

2 2 3 A

ocoopnb *

pnb = —ho ..

2 2

-n oh sh, ch2n3 sch2ch7n3

y

(c02? nb) 2

7 "

40 level K:

ocoopnb

A freshly prepared (H. Davies and M. Schwarz, JOC, 30, 1242 (1965)) solution of p-nitrophenyldiazomethane (29 mmol) in 150 ml ether is stirred with a solution of 1.0 g oxomalonic acid monohydrate (molecular weight = 136 ; 7.35 mmol) in 50 ml of ethyl acetate (EA) at 0 ° C. After 272 hours the yellow solution is concentrated on a rotary evaporator with gentle heating to about half the volume, dried over anhydrous magnesium sulfate, filtered and concentrated to an oil as described above. 3.54 mg of trans-3- (lp-nitrobenzylcarbonyldioxyethyl) -4- [2,2-bis- (2-azidoethyl) -thioethyl] -2- are added to the crude p-nitrobenzyl ester in 50 ml of toluene (Toi.) azetidinone (molecular weight = 524; 6.75 mmol). The reaction mixture is heated with stirring on an oil bath, allowing about 1 h of the toluene to be distilled off. Toluene (dried over a 3A1 / 16 "Linde molecular sieve) is again added to make up the volume to 50 ml and the azeo drying process is repeated three times. The solution is then refluxed under N2 for one hour and the azeo drying process becomes one last time repeated and the reflux treatment is continued for another hour. When the solution obtained is concentrated under a stream of N2, crude is obtained. The raw material is chromatographed on silica gel to give 1. The cis-diastereoisomers or the cis-trans mixture is analogous Way get.

y-

s ch .. ck, n- / 2 2 3

xsch-.ch-n.

-n 0h

2 3

soci.

pnb).

50

55

60

65

3pnb

O

sch0ch_h „- £ -3" \ SCH-; CHcN,

-w «1

Cl

(co, pnb).

ocoopnb i

r sch2ch2n3 sch2ch2n3

H

(c02? nb) 2

-Sy—>

jLnir /

p3p sc. dmf

23

633 800

To a solution of 2.80 g _1 (molecular weight = 912; 3.07 mmol) in 35 ml THF (distilled from lithium aluminum hydride) at - 20 ° C, 0.3 ml pyridine (molecular weight = 79; q = 0.982; 3, 73 mmol) (distilled from NaH and stored over a 4A Linde molecular sieve). While stirring under N2, 0.438 g of thionyl chloride (molecular weight = 119; 3.68 mmol) in 1 ml of THF is added dropwise. The reaction mixture is stirred under N2 for 5 minutes at -20 ° C. and then for 1 hour at 0 ° C. and finally for 1 hour at 25 ° C. The pyridine hydrochloride is filtered off under N2 and washed twice with benzene (dried over a 3A1 / 16 "Linde molecular sieve). The combined filtrates and washing solutions are concentrated under a stream of N2, slurried in a small volume of benzene with anhydrous MgSO4, filtered under N2 and then concentrated under a stream of N2. When pumped under high vacuum for a hour

3rd

0.2 ml of Br2 (molecular weight = 160; 8 = 3.12; 3.9 mmol) are added to 7.8 ml of pentane (dried over a 4A Linde molecular sieve). To a solution of 950 mg 2 (molecular weight = 896; 1.06 mmol) in 15 ml of diethyl ether (dried over a 3A1 / 16 "Linde molecular sieve) is added dropwise at 0 ° C and under N2 with stirring with 2.3 ml of The above 0.40m Br2 solution (1.13 mmol) After stirring for 10 minutes at 0 ° C., 114 μl of cyclohexene (molecular weight = 82.8 = 0.81; 1.13 mmol) are added, after 5 minutes at 0 ° C. 53 mg of 57% NaH (57% of 53 mg = 30.2 mg, molecular weight of 24.1.26 mmol) in mineral oil are added to the stirred reaction mixture, and 14 ml of ice-cold DMF (distilled from anhydrous CaS04 at 40 mm and man To this freshly prepared chlorine compound, 0.885 g triphenylphosphine (mol weight = 262; 3.38 mmol) in 66 ml 9: 1 dimethylformamide (DMF) / H20 and then 550 mg K2HPÖ4 (mol weight = 174; 3.16 mmol) The reaction mixture is stirred for 35 minutes at 25 ° C. After dilution with EA and kitchen salt solution, the layers are separated and the aqueous layer is washed three times with EA extracted. The combined EA layers are washed with brine, dried over anhydrous magnesium sulfate, filtered and concentrated under a stream of N2 to give crude 2. The material is chromatographed on silica gel, giving 2. The cis-di-stereoisomer or the cis-trans mixture is obtained in an analogous manner.

15

Level L:

stored over a 4A Linde molecular sieve). Stirring is continued for 3 hours under N2 at 0 ° C. The reaction mixture is poured onto a stirred ice-cold mixture of 2.5 ml in KH2P04- 40 ml H2ö-75 ml EA. After separating 60 of the layers, the aqueous layer is saturated with NaCl and extracted again with EA. The combined organic layers are extracted once with brine, dried over anhydrous MgSO4, filtered and concentrated under a stream of N2 and then pumped under high vacuum, 65 to give the crude 3. Preparative thin layer chromatography on silica gel gives 3. The cis-diastereoisomers or the cis-trans mixture are obtained in an analogous manner.

ocoopnb

S-CH2CH2N3

7

_n h

Y

(c02? nb) 2

S-CH2CH2N3 • + Br2

ocoopnb

+ I

Br r

sch2ch2n3

sch, ch2n3

(CO, FNS).

ocoopnb?

T

, sch2ch2n3

(c02pn3).

+

633 800

Level M:

24th

OCOOPNB

sch2CH2N3

- + Br0 'AtheT / C5H12 x

£ -J>

(c02pnb),

OCOOPNB

Br

; hch

.he

sch2ch7n3

(C02pnb) 2

+ Nali dmf

->

OCOOPNB Br sch2ch2n3

(C02PNB).

0.2 ml of Br2 (molecular weight = 160; 3.9 mmol) are added to 9.16 ml of pentane (dried over a 4A Linde molecular sieve). To 474 mg 3 (molecular weight = 793; 0.598 mmol) in 13 ml diethyl ether (dried over a 3A1 / 16 "Linde molecular sieve) at 0 ° C. under N2, 1.52 ml of the above-mentioned 0.42 m Br2 solution are added dropwise with stirring (0.63 mmol) After 15 minutes at 0 ° C., 33 mg of 57% NaH (57% of 33 mg = 18.8 mg; molecular weight = 24; 0.78 mmol) are added, and immediately 6 35 ml of ice-cold DMF (distilled from anhydrous CaS04 at 40 mm and stored over a 4A Linde molecular sieve) The reaction mixture is stirred for 1'A hours at 0 ° C., then to a stirred ice-cold mixture of 1.6 ml in KH2P04-20 ml of H20 and 40 are poured into 20 ml of EA. The layers are separated and the aqueous layer is saturated with NaCl and extracted again with further EA. The combined organic layers are washed once with salt water, dried over anhydrous magnesium sulfate and filtered. The filtrate is concentrated. 45 occurs under a stream of N2 and then placed in a high vacuum, being you get the raw 4. Preparative thin-layer chromatography on silica gel gives 4. The cis-diastereoisomers or the cis-trans mixture are obtained in an analogous manner.

50

Stages:

OCOOPNB Br

\

\ - NX

. ~ -sch2CH2N3 + y

OCOOPNB

(c02? n3) 2

DM SO

-sch2ch2n3

(C02PNB) 2

25th

633 800

To 210 mg4 (molecular weight = 871; 0.241 mmol), dissolved in 0.5 ml DMSO (distilled from CaH2 at 8 mm and stored over a 4A Linde molecular sieve), 40 mg l, 5-diazobicyclo- [5, 4.0] -undec-5-ene (distilled at 80 ° C / 2 mm) (molecular weight = 152; 0.263 mmol) in 0.7 ml of dimethyl sulfoxide (DMSO). The solution is stirred for 4 hours under N2 and then added to a stirred ice-cold mixture of 0.48 ml of KH2P04.7 ml of HzO and 10 ml of EA. After separation of the layers, the aqueous layer is extracted again with EA. The combined organic layers are washed once with salt water and then dried over anhydrous magnesium sulfate, filtered and concentrated under a stream of N2 and then placed under a high vacuum to give the crude 5. Preparative thin-layer chromatography on silica gel gives 5. The cis-diastereoisomers or the cis-trans mixture are obtained in an analogous manner.

Level O:

OCOOPNB

the aqueous layer is then extracted two more times with EA. The combined organic layers are extracted with salt water, dried over anhydrous magnesium sulfate, filtered and concentrated under a stream of nitrogen.

s giving raw 6. Preparative thin layer chromatography on silica gel gives 6. The cis-diastereoisomers or the cis-trans mixture is obtained in an analogous manner.

ic level P:

cc0pn3

SCH2CH2N3 + LiI

^ -Colliditi

(c02fne) 2

ocoopnb f

: H2CH2N3

co2pnb

To a solution of 187 mg 5 (molecular weight = 791; 0.236 mmol) in 2.5 ml s-collidine (distilled from powdered KOH at 30 mm pressure) 45 mg of anhydrous LiJ (dried for a few hours at 100 ° C. over P2Os) Vacuum) (molecular weight = 134; 0.336 mmol). The reaction mixture is heated to 120 ° C. on an oil bath while stirring under N2. After a total of 25 minutes, the reaction mixture is cooled to 25 ° C., diluted with CH2C12 and transferred to a round bottom flask in order to concentrate under a stream of N2 and under high vacuum. The residue is divided between 10 ml EA and 1.8 ml in 10 ml H20 in KH2P04 and sch2cr2n3 4-

co2pnb

25th

30th

35

dmso

->

II SCU2 ™ 2n3 c02? Nb

To a solution of the mixed diastereoisomers 6

40 (34 mg; molecular weight = 612; 0.055 mmol) in 0.2 ml DMSO (distilled from CaH2 at 8 mm and stored over a 4A Linde molecular sieve) are added with stirring 9.5 (11.5-diazobicyclo- [5 , 4.0) -undec-5-ene (distilled at - 80 ° C / 2 mm) (molecular weight = 152; e = 1;

45 0.0625 mmol). The solution is stirred under nitrogen for 15 minutes and diluted to a total volume of 1 ml with CHC13 and added immediately to 2 to 1000 silica gel plates. The band of the product shows 7 as a mixture of cis and trans diastereoisomers.

50

Level Q:

OCOQPNB

Oh

-n -

.sch7ch2n3

-co.fnb

JL

H? / 2.2 kg / cr '

y /

-n-

. SCH20H., NK2

• co-, h

7

In the presence of 61 mg of PtO2, 61 mg of 7 (molecular weight = 612; 0.1 mmol) in 6 ml of dioxane, 6 ml of THF, 3 ml of H204 are hydrogenated with hydrogen at a pressure of 2.8 kg / cm2. The reaction mixture is filtered through a Celite filter and washed with 2 ml of 0.1 in pH 7 phosphate buffer. To

8th

After concentrating in vacuo to the cloud point, the aqueous mixture is extracted with ethyl acetate. The aqueous layer is concentrated to a small volume and placed on a column of 100 g XAD-2 resin. When eluting with H20 and discarding the initial fractions, the fraction

633 800

26

nen, which contain the product, freeze-dried, 8 being obtained as a mixture of cis and trans diastereoisomers.

The procedure described below for the enzymatic N-deacylation of thienamycin can be used for all isomers of thienamycin, especially for the pronounced N-acetyl isomers 890A and 890A3, which are described below.

Deacetylation of N-acetylthienamycin

A 1% (w / v) suspension of fertile turf soil is made by adding 1 g of the turf soil to 100 ml of sterile phosphate buffer saline, the phosphate buffer saline having the following composition:

Phosphate buffer saline

NaCl 8.8 g

In phosphate buffer, pH 7.5 + 10 ml distilled water 1000 ml

+ In the phosphate buffer, pH 7.5

16 ml in KH2P04 are mixed with 84 ml in K2HP04. The pH of the phosphate buffer is adjusted to 7.5 by adding a small amount of either Im KH2P04 or Im K2HP04.

Aliquots of this 1% stock earth suspension are prepared for the preparation of 10 X, 100 X and 1000 X dilutions.

1 ml portions of the stock suspension or 1 ml portions of the 10 X, 100 X and 1000 X dilutions are added to 2 ml portions of sterile, 1.0% agar solutions at 48 ° C. The mixture is quickly poured over the surface of sterile petri dishes 85 mm in diameter containing 20 ml of medium A. Medium A has the following composition:

Medium A

KH2P04 3.0 g

K2HP04 7.0 g

MgS04 0.1 g distilled H20 1000 ml

N-acetylethanolamine solution + 8.5 ml + N-acetylethanolamine solution

N-Acetylethanolamine is diluted 10X in HzO and membrane sterilized. This solution is added after the autoclave treatment.

For solid media: add 20 g agar.

The petri dishes are incubated at 28 ° C for 18 days. A well-isolated colony is picked up and spread on a Petri dish containing Medium B. Medium B has the following composition:

Medium B

Tomato paste 40 g ground wheat flour 15 g dest. Water 1000 ml pH: adjusted to pH 6 using NaOH For solid media: add 20 g agar.

A single culture is selected and grown on a medium B agar culture at 28 ° C for 2 days. Part of the growth of this culture is spread onto the surface of 6 cultures made from Medium B. These cultures are incubated at 28 ° C for 2 days. This culture was identified as Protaminobacter ruber and designated MB-3528 in the culture collection by Merck- & Co., Ine, Rahway, N.J., USA and a sample was deposited with the Agricultural Research Service, U.S. Department of Agriculture, Accessions No. NRRLB-8143.

Part of the growth of Protaminobacter ruber MB-3528 agar culture is used to inoculate a 250 ml Erlen-5 Meyer flask containing 50 ml of medium C. Medium C has the following composition:

Medium C

Dextrose 20 g

îoPharmamedia 8 g liquid from soaked corn

(Wet basis) 5 g distilled water 1000 ml pH: adjusted to 7 with NaOH or HCl

15 N-acetylethanolamine solution + 8.5 ml

+ N-acetylethanolamine solution

N-Acetylethanolamine is diluted 10X in H20 and membrane sterilized. This solution is added after the autoclave treatment.

The flask is shaken at 28 ° C and 220 rpm on a vibrator for 4 days. A 25 ml portion from the flask is centrifuged at 8000 rpm for 15 minutes. The supernatant 25 is removed and the cells on the surface of the solid medium are scraped off and added to a 0.5 ml 0.05 M potassium phosphate buffer, pH 7.4. The resulting suspension is sonicated using a Branson Instruments Model LS-75 Sonifier with 30 1/2 inch probe at setting 4 with four 15 second intervals while the suspension is cooled in ice water during and between interruptions becomes. A 10 | xl portion of the sound-treated product is mixed with 25 μl solution, containing 840 (ig / ml N-acetylthienamycin in 10 mmol potassium 35 phosphate buffer, pH 7, and incubated overnight at 28 ° C. Controls, which Antibiotics and buffers alone were included, and sonicated cells and buffers without antibiotics were also made.After incubation overnight at 28 ° C., amounts of 2 ml were placed on cellulose-coated 40 thin-layer chromatography plates which were developed in ethanol: H20, 70: 30. After air drying, the thin layer chromatography plate is placed on a plate infected with Staphylococcus aureus ATCC 6538P for 5 minutes.

45 The infected plates were prepared as follows: The overnight growth of the organism Staphylococcus aureus ATCC 6538P used for the infection in meat broth plus 0.2% yeast extract is diluted to a suspension with meat broth plus 0.2% He-fe extract, which has a 60% transmittance at a wavelength of 660 nm. This suspension is added to Difco nutrient agar supplemented with 2.0 g / 1 Difco yeast extract at 37 to 48 ° C to obtain a composition containing 33.2 ml of the suspension per liter of agar. 40 ml of this suspension are placed on petri dishes,

55 22.5 cm X 22.5 cm, cast and these plates are cooled and kept at 4 ° C until used (5 days maximum).

The thin layer chromatography plate is removed and the plate containing the organism is incubated at 60 35 ° C. overnight. In addition to the unreacted bioactive N-acetylthienamycin with a point at Rf 0.7-0.89, a bioactive point was observed at Rf 0.44-0.47, which is attributed to thienamycin. Control incubation mixtures of the antibiotic plus buffer, sonicated cells plus 65 buffers and antibiotic plus buffers to which sonicated cells had been added shortly before thin layer chromatography application showed no bioactive material at Rf 0.44-0.47.

27

633 800

Preparation of 890A h a special isomer OH

0

-SCH2CH2NHCCK3 COOH

A tube of a freeze-dried culture of Strep-tomyces flavogriseus MA-4434 (NRRL 8139) is opened aseptically and the contents are suspended in a tube containing 0.8 ml of sterile Davis salt of the following composition:

Davis salt sodium citrate k, hpo4 kh2po4 (nh4) 2so4

MgS04 ■ 7H20 distilled water

0.5 g 7.0 g 3.0 g 1.0 g 0.1 g 1000 ml

This suspension is used to incubate four cultures of medium A with the following composition:

Medium A Glycerin primary yeast fish meal distilled water agar

20.0 g 5.0 g 15.0 g 1000 ml 20.0 g pH: adjusted to 7.2 using NaOH

Inoculated cultures are incubated at 27-28 ° C for one week and then stored at 4-6 ° C until use. A 1/3 portion of the growth of three cultures is used to inoculate 9 stoppered Erlenmeyer flasks containing 50 ml of Medium B of the following composition:

Medium B

Yeast autolysate (+ ardamine) glucose

Phosphate buffer ++ MgS04 • 7H20 distilled water

10.0 g 10.0 g 2.0 ml 50 mg 1000 ml pH: adjusted to 6.5 using HCl or NaOH

+ Ardamine: Yeast Products Corporation ++ phosphate buffer solution KH2P04 91.0 g

Na2HP04 95.0 g distilled water 1000 ml

The flasks containing the germs are shaken for 1 day at 27-28 ° C on a vibrator at 220 rpm. The flasks and contents are kept at 4 ° C for 1 day.

33 2-1 ErIenmeyer flasks, each containing 250 ml of medium C, are inoculated with 8 ml per flask of growth from the flask containing the germs. Medium C has the following composition:

Medium C tomato paste primary yeast dextrin (Amidex)

20.0 g 10.0 g 20.0 g

CoCl2 • 6H20 5.0 mg distilled water 1000 ml pH: adjusted to 7.2-7.4 by NaOH

s After the inoculation, the production pistons are at

24 ° C incubated with shaking on a vibrator that is operated at 212 rpm for 4 days. The flasks are then emptied and the content checked for activity using standard Salmonella gallinarum ioMB1287 and Vibrio percolans ATCC 8461 plates using 1.25 cm disks immersed in the centrifuged broth samples. If necessary, the samples are diluted with 0.02m phosphate buffer, pH 7.0. The results are given below:

15

Harvest Age hours h 96 pH 7.2

Salmonella gallinarum (mm zone) 29.5

Vibrio percolans 1/10 dilution (mm zone) 31 20 890 test units 40

7 1 of the entire fermentation broth are cooled to 3 ° C. and centrifuged in 200 ml portions at 9000 rpm for 15 minutes.

25 To the combined supernatant liquids are added 7 ml 0.1m neutral EDTA and the whole sample is placed on a Dowex-1 X 2 (Cl-), 50-100 mesh column with bed dimensions of 5.1 X 25 cm and one Flow rate of 40 ml / min. The column is washed with 500 ml of deionized water, containing 5 ml in Tris-HCl buffer, pH 7.0, and

25 [imol neutral EDTA, washed. The antibiotic is eluted with 1 liter of deionized water containing 50 g of sodium chloride, and the column is then washed with 300 ml of deionized water. 100 ml fractions are

35 melts, starting from the first appearance of salt at the column exit. The bioactivity is present in fractions 1 to 10 with a peak in fraction 2. Fractions 2 to 5, which contain 17% of the activity used, are pooled.

40 The combined fractions are concentrated to 110 ml by means of a rotary evaporator under reduced pressure and the pH is adjusted to 5.8 by adding 4.2 ml in HCl. The concentrate set in this way is placed on a column of XAD-2 with a bed dimension of 45 3.8 × 50 cm, which had previously been washed with 3 160% aqueous acetone (volume / volume) and then with 3 l of deionized water, 3 1 of 5% (weight / volume) sodium chloride and 1125% (weight / volume) sodium chloride in deionized water. The concentrate used is allowed to flow down to the level of the bed. The antibiotic is eluted with deionized water at a flow rate of 15 ml / min. 22 fractions of 100 ml each are collected, calculated from the first application of the sample. The bioactivity appears in fractions 55 to 6 with a peak in fractions 9 and 10. Fractions 9 to 20 are combined for further processing. Similar fractions are pooled and the pooled fractions are concentrated to 70 ml using a rotary evaporator under reduced pressure.

60

The concentrate is placed on a Dowex-1 X 4 (Cl -) 400 mesh column with a bed dimension of 2.2 X 27 cm. The column is washed with 50 ml deionized water and the antibiotic eluted with 2 10.070m NaCl + 0.005m 65 NH4C1 + 0.0001m NH3 in deionized water at a flow rate of 2 ml / min. 9.5 ml fractions are collected.

The main peak of the antibiotic is eluted in the fracture

633 800

28

ions 142 to 163. Fractions 146 to 157 are combined for further processing.

The combined fractions 146-157 are concentrated to 3 ml by evaporation under reduced pressure on a rotary evaporator and the concentrate is brought to pH 6.5 by adding 5 ml of NH 3. The concentrate is placed on a column (2.2 X 70 cm) of a Bio-Gel P-2 (200 to 400 mesh), which had been washed beforehand with 20 ml of 5M NaCl and 500 ml of deionized water. After the concentrate had run down to bed level, 2 rinses with 1 ml of deionized water each were carried out and again run down to bed level. The column is then eluted with deionized water at a flow rate of 0.6 ml / min. 2.9 ml fractions are collected.

The main peak of the antibiotic is eluted in fractions 63 to 70. Fractions 64 and 65 are pooled for further processing. The combined fractions 64 and 65 are concentrated on a thin-layer evaporator to 2 ml under reduced pressure and then frozen in an envelope and lyophilized for 8 hours in a 14 ml screw-capped vessel to give 2.25 mg of the essentially pure 890At antibiotic . The N-acetyl group is cleaved off as described previously, giving the free base:

f

/ \ / V-SCH0CH0KH ..

ì N ÌLCOOH

0

Production of 890A3, a special isomer

0

SCH2CH2MHCCH.

A tube of a lyophilized culture of Streptomyces flavogriseus MA-4434 (NRRL 8139) is opened aseptically and the contents are suspended in a tube containing 0.8 ml of a sterile Davis saline solution of the following composition:

Davis salt

Sodium citrate 0.5 g

K2HP04 7.0 g

KH2P04 3.0 g

(NH4) 2S04 1.0 g

MgS04-7H20 0.1g distilled water 1000 ml

This suspension is used to inoculate 4 cultures of medium A with the following composition:

Medium A

Glycerin 20.0 g primary yeast 5.0 g

Fish meal 15.0g distilled water 1000 ml

Agar 20.0 g pH: adjusted to 7.2 under NaOH

The inoculated cultures are incubated at 27-28 ° C for 1 week and then stored at 4-6 ° C until use (no longer than 21 days).

A 1/3 portion of the growth from 4 cultures is used to inoculate 12 blocked 250 ml Erlenmeyer flasks containing 50 ml medium B of the following composition:

Medium B îo yeast autolysate (+ Ardamine) 10.0 g glucose 10.0 g

Phosphate buffer ++ 2.0 ml

MgS04 • 7H20 50 mg distilled H20 1000 ml

15 pH: adjusted to 6.5 under HCl or NaOH + Ardamine: Yeast Products Corporation ++ phosphate buffer solution KH2P04 91.0 g

Na2HP04 95.0 g

20 distilled water 1000 ml

The bottle with the germs is shaken for 1 day at 27-28 ° C on a vibrating machine at 220 rpm. The bottle and its contents are kept stationary at 4 ° C for 1 day. 25 44 2-1 Erlenmeyer flasks, each containing 200 ml of medium C, are mixed with 8 ml per flask of growth from the inoculated bottles. Medium C has the following composition:

20.0 g 10.0 g 20.0 g

5.0 mg 1000 ml -7.4 with NaOH

After the inoculation, the production flasks are shaken at 4o 24 ° C for 4 days and 5 hours on a shaking machine that is operated at 212 rpm. The flasks are then emptied and assayed for activity using standard Salmonella gallinarum MB1287 and Vibro percolans ATCC 8461 test plates using 1.25 cm Pro-45 which are immersed in the centrifuged samples of the broth. The samples are diluted with 0.2m phosphate buffer, pH 7.0, if necessary. The results are given below:

so time of harvest (Harvest

Age hours) h 101

pH 7.2

Salmonella gallinarum (mm zone) 35 Vibrio percolans 1/10 dilution

55 (mm zone) 34

890 sample units 121

A total of 7 l of the entire broth are obtained from this fermentation and these are cooled to 3 ° C. and centrifuged in 200 ml portions at 9000 rpm for 15 minutes each. 1.7 ml of 0.1 m neutral EDTA are added to the combined supernatant solutions and the mixture is kept at 3 ° C.

The above fermentation is repeated under identical conditions with the exception that the 44 2-1 ErIen-meyer flasks are inoculated with 7 ml per bottle of growth from the bottles containing the cultures; the pH and the test results obtained are given below:

30 medium C tomato paste primary yeast dextrin (Amidex)

COCl2 • 6H20 35 distilled water pH: adjusted to 7.2

29

633 800

Time of harvest (Harvest

Age hours) h

101

pH

7.3

Salmonella gallinarum (mm zone)

38

Vibrio percolans 1/10 dilution

(mm zone)

39

890 sample units

92.8

A total of 7.41 of the entire broth obtained from this fermentation are cooled to 3 ° C. and centrifuged for 15 minutes in 200 ml portions at 9000 rpm. 1.8 ml of 0.1 m neutral EDTA are added to the combined supernatant solutions.

The supernatant from the centrifuged broths as obtained from the above two fermentations in this example is combined to give a total volume of 13 liters.

The combined supernatant solutions are passed through a column of Dowex-1 X 2 (Cl-), 50-100 meshes with a bed dimension of 4.7 cm X 50 cm and a flow rate of 60 ml / min. The column is washed with 11 deionized water and the antibiotic is eluted with 5 l of a 5% (weight / volume) NaCl solution containing 0.01 M Tris-HCl buffer, pH 7.0, and 25 [mol EDTA. 220 ml fractions are collected at a flow rate of 50 ml / min and the fractions are checked against Salmonella gallinarum MB 1287 plates.

Antibiotic activity is present in fractions 3 to 26 with a peak in fractions 5 and 6. Fractions 5 to 9 are combined for further processing. The pH of the pooled fractions is 10.8 and the pooled fractions contain 24% of the initial bioactivity as measured against Salmonella gallinarum MB1287 plates.

The combined fractions 5 to 9 are concentrated to 150 ml by means of a rotary evaporator under reduced pressure and placed on a column (4.9 cm X 47 cm) of XAD-2, which had been previously washed with 5 160% (volume / volume ) aqueous acetone and then with 5 1 of deionized water and 5 150 g / 1 NaCl in deionized water. The sample is applied in 20 ml increments, soaking the column to bed level each time. When application is complete, three 20 ml portions of deionized water are added and each time allowed to flow to bed level. The sample is eluted with deionized water at a flow rate of 20 ml / min. All measures using the XAD column are carried out at room temperature (24 ° C) and the eluted fractions are quickly cooled in an ice bath immediately after collection. Fractions from 95 to 230 ml are collected.

The antibiotic activity appears in fractions 2 to 21, as measured from a sample of Salmonella gallinarum MB1287 plates, with a peak in fractions 5 to 7 (which make up 510 to 895 ml of eluted volume, calculated from the first application of the deionized water). Fractions 6 to 21 are pooled.

The pooled fractions 6 to 21 are concentrated to 68 ml on a rotary evaporator under reduced pressure and then diluted to 112 ml by adding deionized water. The concentrate is placed on a column (2.2 cm X 21 cm) of Dowex-1 X 4 (CI ~) 400 mesh. The column is washed with 20 ml deionized water and the antibiotic is eluted with 210.07m NaClH-0.005m

NH4C1 + 0.0001m NH3 in deionized water with a flow rate of 1.6 ml / min. Fractions of 8 ml each are collected. Fractions 157 to 179 are the most active and are pooled.

5 These combined fractions are concentrated on a rotary evaporator under reduced pressure to 7 ml, the pH is adjusted to 7.5 by adding 20 | xl in NaOH. The solution is further concentrated to 5 ml and placed on a column (2.2 X 75 cm) of Bio-Gel P-2,200 to ic 400 mesh. The sample is washed in the column bed with two washes of 1 ml of deionized water each and eluted with deionized water at a flow rate of 0.6 ml / min. Ten fractions of 3.3 ml and then 60 fractions of 2.65 ml and ten fractions of 15 2.0 ml are collected.

The fractions are adjusted to the pH range between 7.5 and 8.0 by adding 1.5 to 2.5 | d 0.1m NaOH. Fractions 62, 63, 64 and 65 are frozen and individually lyophilized in 14 ml glass tubes and stored at -20 ° C under a vacuum.

To isolate the antibiotic 890A3 and 890AJ, one takes advantage of the relatively higher resistance of the antibiotic 890A3 to degradation by penicillinase in the following manner:

25 Bio-Gel fraction 63 is combined with fractions 61, 66 and 67 from the Bio-Gel column. 0.2 ml in Tris-HCl buffer, pH 7.5 and 0.2 ml penicillinase (Difco "Bacto-Penase") are added to these four combined fractions. After 113 minutes at 23 ° C, another 0.2 ml of penicillinase are added. After a further 7 hours at room temperature, a further 0.2 ml of penicillinase are added, and after a further 2 hours at room temperature the reaction mixture is cooled on an ice bath. The finished reaction mixture is diluted to 15 ml by adding 5 ml of deionized water. 35 The reaction mixture is adsorbed on a Dowex-1 X 4 (Cl-) 400 mesh column, bed dimension 2.15 X 40 cm. The antibiotic 890A3 is eluted with 0.07m NaCl + 0.005m NH4CI + 0.0001m NH3 in deionized water at a flow rate of 3 ml / min. Fractions of 13.8 ml 40 are collected in each case. Fractions 187 to 200 are pooled.

The combined fractions are concentrated to 4 ml using a rotary evaporator under reduced pressure and the concentrate is applied to a column (2.2 × 70 cm) Bio-Gel P-2, 45 200-400 meshes. Antibiotic 890A3 is eluted with deionized water at a flow rate of 0.6 ml / min. 30 fractions of 3.3 ml and then 50 fractions of 2.65 ml are collected.

Fractions 66 to 70 are combined and the combined samples are concentrated to 1.5 ml under reduced pressure using a rotary evaporator and the concentrate is frozen and freeze-dried to give 5.4 mg of a solid containing the 890A3 antibiotic and residual salt . The N-acetyl group is split off, as previously described, 55 so that the free base is obtained:

60

CH2CH2NH2 cooh c

Claims (6)

633 800 2nd PATENT CLAIMS 1. Process for the preparation of guanidines of the general formula (II) (II) sh, ch ^ n = c-x 2 ^ t cooh y 10th wherein Y is -NR'R2 and X-NR'R2, where R! and R2 are independently selected from hydrogen, nitro, hydroxyl, alkoxyl with 1 to 6 carbon atoms, amino, mono-, di- and trialkylamino, in which the alkyl groups each contain 1 to 6 carbon atoms; R1 and R2 can be linked together to form a substituted or unsubstituted mono- or dicyclic heteroaryl or heterocyclyl containing (together with the nitrogen atom to which they are attached) 4 to 10 atoms, one or more of which represent a further heteroatom the group can be oxygen, sulfur or nitrogen; R1 and R2 are substituted or unsubstituted, namely: cyano; Hydrogen; Carbamoyl; Carboxyl; Alkoxycarbonyl and alkyl of 1 to 10 carbon atoms; Alkenyl of 2 to 10 carbon atoms; Alkynyl of 2 to 10 carbon atoms; Cycloalkyl of 3 to 10 carbon atoms; Cycloalkylalkyl and cycloalkylalkenyl of 4 to 12 carbon atoms; Cycloalkenyl, cycloalkenylalkenyl and cycloalkenylalkyl of 3 to 10.4 to 12 and 4 to 12 carbon atoms, respectively; Aryl of 6 to 10 carbon atoms, aralkyl, ar alkenyl and aralkynyl of 7 to 16 carbon atoms; mono- and bicyclic heteroaryl and heteroaralkyl having 4 to 10 ring atoms, one or more of which is a heteroatom from the group consisting of oxygen, sulfur or nitrogen, and in which the alkyl group of the heteroaralkyl radical contains 1 to 6 carbon atoms; mono- and bicyclic heterocyclyl and heterocyclyl alkyl having 4 to 10 ring atoms, one or more of which is a heteroatom from the group consisting of oxygen, sulfur or nitrogen, and in which the alkyl group of the heterocyclylalkyl radical contains 1 to 6 carbon atoms; and wherein the aforementioned substituent or substituents on R1, R2 or on the ring formed by the connection of R1 and R2 are selected from the group consisting of: chlorine, bromine, iodine and fluorine; Azido; Alkyl with 1 to 4 carbon atoms; Thio; Sulpho; Phosphono; Cyanothio (-SCN); Nitro; Cyano; Amino; Hydrazino; mono-, di and trialkyl-substituted amino and hydrazino, wherein the alkyl has 1 to 6 carbon atoms; Hydroxyl; Alkoxyl of 1 to 6 carbon atoms; Alkylthio with 1 to 6 carbon atoms; Carboxyl, oxo; Alkoxycarbonyl having 1 to 6 carbon atoms in the alkoxy group; Acyloxy with 2 to 10 carbon atoms; Carbamoyl and mono- and dialkylcarbamoyl, in which the alkyl group each has 1 to 4 carbon atoms, and their pharmaceutically acceptable salts, characterized in that thienamycin (I) 15 25th 30th wherein X "O or S and -X" R ° is the leaving group and R1 and R2 are as defined above and R ° is any residue, treated and optionally obtained compounds converted into their pharmaceutically acceptable salts. 2. The method according to claim 1, characterized in that R1 and R2 are independently selected from the group consisting of: hydrogen; substituted and unsubstituted radicals, namely: alkyl of 1 to 6 carbon atoms; Alkenyl of 3 to 6 carbon atoms; Cycloalkyl, cycloalkylalkyl, cycloalkenyl and cycloalkenylalkyl having 3 to 6, 4 to 7.4 to 6 or 4 to 7 carbon atoms; Aralkyl and aralkenyl of 7 to 10 carbon atoms; and monocyclic heteroaralkyl with 5 to 6 ring atoms, one or more of which are selected from the group: contain oxygen, sulfur and nitrogen and 1 to 3 carbon atoms in the alkyl group; wherein the ring or chain substituent or substituents are selected from the group consisting of chlorine, fluorine, hydroxyl, alkoxyl with 1 to 3 carbon atoms, dialkylamino with 1 to 3 carbon atoms in each alkyl group and alkylthio with respect to the definition of R1 and R2 with 1 to 3 carbon atoms. 3. The method according to claim 1, characterized in that R1 and R2 are independently selected from the group consisting of: hydrogen; Alkyl; Alkenyl; Nitro; Amino and phenyl. 4. The method according to claim 1, characterized in that R1 and R2 are selected from the group consisting of hydrogen, alkyl having 1 to 6 carbon atoms and alkenyl having 2 to 6 carbon atoms. 5. The method according to claim 1, characterized in that a compound of the formula oh <f -N- -sch2ck2n cooh nh_, / 2nd = c XNH_ 40 manufactures. 50 55 ok k _N. sch2ch2nh2 . cooh (I) 60