Note: Descriptions are shown in the official language in which they were submitted.
The present invention relates to novel 3~-amino
or 3~-acylamino-4a-acetoxyazetidinone compounds which are
useful antibacterials for controlling ~-lactamase producing
gram-negative bacteria and other pathogens, or are useful
intermediates in the preparation of said antibiotics, and to
a novel process for the preparation of said compounds.
This invention relates to ~-lactam antibiotics.
In particular, it relates to monocyclic ~-lactam antibiotics
which are 3~-acylaminoazetidin-2-ones.
The penicillins and cephalosporins are well known
~~lactam antibiotics which are bicyclic compounds containing
a fused ring system. The penicillins have the 4-membered
~-lactam ring fused to a thiazolidine ring while in the
cephalosporins, the ~-lactam ring is fused to a dihydro-
thiazine ring. Monocyclic ~-lactam antibacterial compounds
are less well known. The monocyclic ~-lactam antibiotic,
nocardicin, has been recently discovered and is described in
Belgium Patent No. 830,934 and by H. Aoki, et al., 15th
Interscience Conference on Antimicrobial Agents and Chemo-
therapy, Abst. No. 97, Sept. 197S.
Nocardicin has the following structural formula.
O H
HOOC CH CHz CHz O ~ _o/ IC~ C N I I ~
OH COOH
In view of the importance of ~-lactam antibiotics
in the treatment of infectious diseases, considerable effort
is directed by microbiologists and chemists to the discovery
X-4683A -2-
and development of other ~-lactam antibiotics which possess
activity against a broader spectrum of microorganisms or
which are more effective than the currently available
antibiotics.
Monocyclic ~-lactam azetidinone compounds having
an acetoxy group in the 4-position previously were unknown.
The present invention provides a process f~r ~re?aring
a 4~-acetoxyazetidin-2-one compound of the formula
O
~0 C-CH3
COOR1 a
wherein R is an amino group or an acylanuno group; of the formula
O H
Il I
R ' -C-N-
wherein R' is Cl-C4 alkyl, cyanomethyl, bromomethyl, chloro-
methyl, phenyl, or a group of the formula
O Z
Ra-O-C-CH-CHz-CHz-O-~ C-
NH
Rb
wherein Ra is hydrogen, benzyl, diphenylmethyl or 4-
methoxybenzyl;
Rb is hydrogen or an amino-protecting group of the
formula
c 11
R -O-C-
wherein Rc is t-butyl, 2,2,2-trichloroethyl, benzyl,
4-nitrobenzyl, cyclopentyl, or cyclohexyl;
3-
6:1~
Z is =O or =N-OZ', wherein Z' is hydrogen, acetyl,
chloroacetyl, triphenylmethyl or p-methoxybenzyl;
or R' is a group of the formula
R''-CH2-
wherein R'' is a phenyl group of the formula
b
b~
wherein b and b' independently are hydrogen, halogen,
hydroxy, Cl-C4 alkyl, C1-C4 alkoxy, amino or amino-
methyl; or R" is thienyl, furyl, thiazolyl, oxazolyl,
isothiazolyl, tetrazolyl, or an isoxazolyl group of the
formula
~/- r~
wherein d is hydrogen, methyl, or a group of the ~:
formula
b'
wherein b and b' have the same meanings defined above,
and d' is hydrogen, methyl, or chloro;
or R' is a phenoxymethyl group of the formula
~ ~ ~ CH2-
b~
wherein b and b' have the same meanings as defined
above;
-4- ~.
i61~
or R' is a group of the formula
R'''-S-CH2-
wherein R''' is a group of the formula
b
b~ ,
wherein b and b' are as defined above,
4-pyridyl, thiazolyl, thiadiazolyl, or oxadiazolyl;
or R' is a group of the formula
H
R''''-C-
Q
wherein R'' " is a phenyl group of the formula
b
b' ~
wherein b and b' are as defined above,
thienyl or furyl;
and Q is amino, hydroxy, carboxy, -SO3H, or -NH-SO3H,
or any of these groups in protected form;
or R' is a group of the formula
R''''-C-
N
o
Z''
wherein R'''' has the same meanings as defined above
and Z'' is hydrogen, acetyl or methyl;
or R' is a group of the formula
H
R''''-C-
N-H
C=O
~ :
.. .,~ -5-
" ~ :
6~
wherein R" " is as defined above: and
Y is a dimethylureido group of the formula
CH3 O
~ 11
- N - C - N CH3
an imidazolidin-2-one group
_Nt\ tN_y,
1 0 o
wherein Y' is hydrogen, Cl-C4 alkyl, C~-C4 alkanoyl,
or methanesulfonyl;
or an N-methylacyl group of the formula
CH3 o
I
- N - C - Y''
wherein Y'' is Cl-C4 alkyl, or a group of the
formula
--~CH =CH)n --~
--J a"
wherein n =O or 1 and a'' is hydrogen,
nitro, or chloro;
Rl is hydrogen or a carboxylic acid protecting group
and a and a' independently are hydrogen, halogen,
hydroxy, protected hydroxy, Cl-C4 alkoxy, amino, protected amino,
amino-methyl or protected amino methyl; which comprises heating
a thiazolidine azetidinone compound of the formula
5a
~ .
. :
6~
O \~/
Ra-C - N
COOR1 a
wherein Ra is Cl-C3 alkyl, phenyl, benzyl or phenoxymethyl;
and Rl, a and a' are as defined above;
with mercuric acetate in the presence of acetic acid to form
a N-propenyl 4a~acetoxyazetidin-2-one amide of the formula
CH3
O C=CH2 ll
Il I /O-C-CH3
COOR1 a
wherein Ra, Rl, a and a' are as defined above; hydrolyzing
the 4~-acetoxyazetidin-2-one amide to form a 4~-acetoxy-
azetidin-2-one of the formula
o
O ~I
Il O-C-CHs
~ C
COORl
wherein Ra, Rl, a and a' are as defined above;
if desired cleaving by conventional methods the 3-position
acyl group to provide a compound of formula I wherein R is
amino; if desired reacylating the 3-amino-4-azetidin-2-
one compound so obtained by conventional methods; and
optionally removing the carboxy, hydroxy or amino protecting
-Sb- ¦
6il ~
groups; and where desired, when Rl is hydrogen, forming
a pharmaceutically acceptable acid addition salt of the
compound of Formula I so produced.
The present invention also provides 4~-
acetoxyazetidin-2-one-compounds of Formula I as
hereinbefore defined; and when Rl is hydrogen, the
pharmaceutically acceptable non-toxic salts thereof.
As previously indicated, when in the above
formula R is an acylamino group, it preferably is a
group of the formula
O H
Il l
R'-C-N-
wherein R' is Cl-C4 alkyl, cyanomethyl, bromomethyl, chloro-
methyl, phenyl, or a group of the formula
O Z
Ra-O-C-CH-CH2-CH2-O--~ ~C-
NH
Rb
wherein Ra is hydrogen, benzyl, diphenylmethyl or 4-methoxy-
benzyl;
Rb is hydrogen or an amino-protecting group of the
formula
o
RC-O-C-
wherein Rc is t-butyl, 2,2,2-trichloroethyl, benzyl,
4-nitrobenzyl, cyclopentyl, or cyclohexyl;
z is =O or =N-OZ', wherein Z' is hydrogen, acetyl,
chloroacetyl, triphenylmethyl or p-methoxybenzyl;
or R' is a group of the formula
R''-CH2-
-5c-
. - ~
1~ 6~3
wherein R'' is a phenyl group of the formula
b~
b~ ==-
wherein b and b' independently are hydrogen, halogen,
hydroxy, Cl-C4 alkyl, Cl-C4 alkoxy, amino or amino-
methyl; or R'' is thienyl, furyl, thiazolyl, oxazolyl,
isothiazolyl, tetrazolyl, or an isoxazolyl group of the
formula
~ /-- or
wherein d is hydrogen, methyl, or a group of the
formula
b
b~
wherein b and b' have the same meanings defined above,
and d' is hydrogen, methyl, or chloro;
or R' is a phenoxymethyl group of the formula
~ ~ -O-CH2-
b~
wherein b and b' have the same meanings as defined
above;
or R' is a group of the formula
R'''-S-CH2-
X-4683A -6-
wherein R''' is a <Jroup Or thc formula
b~ '
wherein b and b' are as defined above,
4-pyridyl, thiazolyl, thiadiazolyl, or oxadiazolyl;
or R' is a group of the formula
H
R ' ' ' ' -C-
Q
wherein R'''' is a phenyl group of the formula
b~
wherein b and b' are as defined above,
thienyl or furyl;
and Q is amino, hydroxy, carboxy, -SO3H, or -NH-SO3H,
or any of these groups in protected form;
or R' is a group of the formula
R ' ' ' '-C-
O
Z
wherein R'''' has the same meanings as defined above
and Z'' is hydrogen, acetyl or methyl; .- -
or R' is a group of the formula
X-4683A -7-
R ' ' ' ' -C-
N-ll
C=O
. y .
wherein R'''' is as defined above and
Y is a dimethylureido group of the formula
CH3 0 H
11
- N - C - N - CH3
an imidazolidin-2-one group
y,
O '.
wherein Y' is hydrogen, Cl-C4 alkyl, C2-C4 alkanoyl
or methanesulfonyl;
or an N-methylacyl group of the formula
CH3 0
Il
- N - C- Y''
wherein Y'' is Cl-C4 alkyl, or a group of the
formula
n \~ ~Xa,,
wherein n = 0 or 1 and a'' is hydrogen,
nitro, or chloro.
X-4683A -8-
The phenyl group represented in the above defi-
nition by the formula
a
a~ ~==-
represents phenyl, 4-hydroxyphenyl, 4-(tetrahydropyran-2-
yloxy)phenyl, 4-benzyloxyphenyl, 3-hydroxyphenyl, 3,4-
dihydroxyphenyl, 4-chlorophenyl, 3,4-dichlorophenyl, 2,6-
dichlorophenyl, 3-bromophenyl, 3-chloro-4-hydroxyphenyl,
3-chloro-4-methylphenyl, 4-t-butylphenyl, 3,4-dimethyl-
phenyl, 2,4-dimethylphenyl, 4-ethylphenyl, 3-methyl-4-
hydroxyphenyl, 4-aminophenyl, 3-aminophenyl, 3-amino-4-
methylphenyl, 2-aminomethylphenyl, 4-aminomethylphenyl, 4-
methoxyphenyl, 2,6-dimethoxyphenyl, 3-ethoxy-4-hydroxy-
phenyl, 4-isopropoxyphenyl, 4-chloro-2-aminomethylphenyl, 3-
bromo-4-methoxyphenyl, 3-methyl-4-aminophenyl, and like
substituted phenyl groups.
The phenyl group represented in the above formula
by
b~
represents phenyl, 4-chlorophenyl, 3,4-dichlorophenyl,
3-bromophenyl, 3-chloro-4-hydroxyphenyl, 4-hydroxyphenyl,
3-hydroxyphenyl, 3,4-dihydroxyphenyl, 4-methylphenyl,
4-t-butylphenyl, 4-ethylphenyl, 4-methoxyphenyl, 3-ethoxy-
4-hydroxyphenyl, 4-aminophenyl, 4-aminomethylphenyl, 3-methyl-
4-aminophenyl, 2,6-dimethoxyphenyl, 3-bromo-4-methoxyphenyl
and like substituted phenyl groups.
X-4683A -9-
Illustrative of the amino-protecting groups defined
in the above formula I by the formula RC-O-C- are t-butyl-
oxycarbonyl, 2,2,2-trichloroethoxycarbonyl, benzyloxycarbonyl,
p-nitrobenzyloxycarbonyl, cyclopentyloxycarbonyl and cyclo-
hexyloxycarbonyl~
As used herein the term thienyl refers to both 2-
thienyl and 3-thienyl; thiazolyl refers to 2-thiazolyl and
5-thiazolyl; tetrazolyl refers to 1- and 2-tetrazolyl;
thiadiazolyl refers to 1,3,4-, 1,2,5- and 1,2,4-thiadiazol-
5-yl; and oxadiazolyl refers to 1,3,4-oxadiazolyl.
Examples of isoxazolyl groups represented in the
above formula I by
\ ~ ~~0/ or
are 3-methylisoxazol-5-yl, 3,4-dimethylisoxazol-5-yl, 4-
chloroisoxazol-5-yl, 3-phenyl-4-chloroisoxazol-5-yl, 3-
(4-chlorophenyl)isoxazol-5-yl, 3-(2-chlorophenyl)isoxazol-
5-yl, 3-(2-chlorophenyl)-4-methylisoxazol-5-yl, 3-(2,6-
20 dichlorophenyl)isoxazol-5-yl, 3-(2,6-dichlorophenyl)-4-
methylisoxazol-5-yl, isoxazole-5-yl, 3-(2,4-dimethyl-
phenyl)-4-methylisoxazol-5-yl, 3-(2,6-dimethylphenyl)-
4-chloroisoxazol-5-yl, 3-(4-methoxyphenyl)isoxazol-5-yl,
3-(2,6-dimethoxyphenyl)-4-chloroisoxazol-S-yl, 3-(3-
chloro-4-hydroxyphenyl)-4-methylisoxazol-5-yl, 3-methyl-5-
chloroisoxazol-4-yl, 3,5-dimethylisoxazol-4-yl, 3-phenyl-
isoxazol-4-yl, 3-(4-chlorophenyl)-5-methylisoxazol-4-yl,
3-(2-chlorophenyl)-5-chloroisoxazol-4-yl, 3-(2,6-dimethoxy-
phenyl)-5-methylisoxazol-4-yl and like isoxazoles.
X-4683A -10-
The compounds of the formula I can be charac-
terized as monocyclic ~-lactam antibiotics or as azetidin-
2-ones which are substituted in the l-position with an ~-
carboxybenzyl or ~-carboxy-substituted benzyl group and in
the 3-position by a ~-amino or ~-acylamino group and which
also are substituted in the 4-position with an actoxy group.
The thiazolidine azetidinone starting material
represented by the formula II is prepared according to the
following method. L-cysteine is reacted with dry acetone at
the reflux temperature to provide 2,2-dimethyl-4-thiazoli-
dinecarboxylic acid. The product is acylated with an acyl
chloride in the presence of propylene oxide to provide
3-acyl-2,2-dimethyl-4-thiazolidinecarboxylic acid repre-
sented by the following formula.
O CH3\~CH3
Ra-C - -N
COOH
wherein Ra is as defined above.
The thiazolidinecarboxylic acid is coupled with an ester of
a phenylglycine represented by the formula,
H
H~N-C - ~ ~
COOR1 a
wherein a, a', and Rl are as defined herein to form the
amide represented by the following formula
~-4G83A -11-
H3C\ /CH3
Ra-~-N ~ ,
~____ ,
~H ~ a
OOR1
The coupling reaction is carried out by first preparing the
active ester of the thiazolidinecarboxylic acid formed with
l-hydroxybenzotriazole (HBT). The active ester then is
reacted with the phenylglycine ester in the presence of
dicyclohexylcarbodiimide.
Phenylglycine esters which can be used to form the
above amides include for example, benzyl phenylglycinate, 4-
methoxybenzyl phenylglycinate, benzyl 4-methoxyphenyl-
glycinate, benzyl 4-chlorophenylglycinate, benzyl 3,4-
dimethylphenylglycinate, benzyl 4-hydroxyphenylglycinate, - ;
2,2,2-trichloroethyl phenylglycinate, 4-methoxybenzyl
3-chloro-4-hydroxyphenylglycinate, benzyl 4-t-butylphenyl-
glycinate, benzyl 4-aminophenylglycinate, and 2,2,2-tri-
chloroethyl 3,4-dichlorophenylglycinate. When the phenyl-
glycine ester is substituted with a reactive functional
group which can interfere with the desired coupling reaction
(N-acylation), for example, a phenolic OH group or an amino
group, such groups are blocked during the amidation reaction
and in subsequent reactions in the process. The amino group
can be blocked with the t-butyloxycarbonyl (t-BOC) group or
the benzyloxycarbonyl group. A phenolic hydroxy substituent
is best blocked with the benzyl group or tetrahydropyran
group.
X-4683A -12-
The thiazolidine amide of the above formula is
then heated at the reflux temperature in a hydrocarbon
solvent such as benzene or toluene with benzoyl peroxide to
form the 5-benzoate derivative represented by the formula
CH3\ /CH3
O-C-~
~OOR- a'
The benzoate is reacted in methylene chloride at about 0C.
with hydrogen chloride to form the corresponding 5~-
chloro compound by replacement of the benzoate function.
The 5~-chloro amide is reacted with sodium hydride under
anhydrous conditions in a halogenated hydrocarbon solvent,
such as methylene chloride, at a temperature between about
0C. and about 30C. to effect an intramolecular cyclization
and provide the substituted thiazolidine azetidin-2-one
represented by the following formula.
H3C\ /CH3
Ra-~-N
_c~
COOR1 a
For convenience the compounds of the above formula
are rc~erred to hcrein as thiazolidine azetidinone deriva-
tivcs. lrhcsc compounds formally are named 2-acyl-3,3-
dialkyl-7-oxo-~-[substituted phenyl]-4-thia-2,6-diazabicyclo-
[3.2.0]heptane-6-acetic acid esters.
X-4683A -13-
f~61i3
The compouncls of formula I are prepared by the
rocc.ss o~lt: l inc(l above 'l'hc rcaction of thc thia~olidine
a~e~idi norlc with mercuric acctatc is carried out in aceti~
acid or in an inert co-solvent with acetic acid. Co-solvents
such as tetrahydrofuran, dioxane, dimethylformamide, or
dimethylacetamide can be used. A large excess of acetic
acid is employed while between 1 mole and 3 moles of
mercuric acetate per mole of the thiazolidine azetidinone is
used. Preferably, 1.5 to 2.0 moles of mercuric acetate per
mole of starting material is employed.
The reaction preferably is carried out in acetic
acid with heating at a temperature between 25 and 75C.
The reaction mixture is filtered after the reaction is
complete to remove insoluble mercury compounds and the
filtrate is evaporated. The product is then extracted from
the residue with a water immiscible organic solvent such as
ethyl acetate. The product, a N-propenylamide of the fore-
going formula III, need not be purified for its hydrolysis
to the amide. The hydrolysis is best carried out between
15 and 55C. in a water-miscible solvent, for example,
tetrahydrofuran, with dilute hydrochloric acid, for example,
between 2 percent and 10 percent hydrochloric acid. Dilute
sulfuric or dilute phosphoric acid also can be used.
Alternatively, the hydrolysis is carried out in aqueous
tetrahydrofuran, preferably 50% aqueous tetrahydrofuran,
with mercuric acetate. Preferably an amount of mercuric
acctatc cqual in weight to the amount of N-propenylamine is
employed. The hydrolysis can be carried out at a tempera-
ture between 15C. and 45C. and preferably at 20C. to 25C.
X-4683A -14-
6~
~ fter the hydrolysis of the N-propenylamide is
complete, thc reaction mixture is evaporated and the residue
containing the product is dissolved in a water immiscible
solvent such as ethyl acetate and the solution is washed
with a dilute base such as sodium bicarbonate to remove
traces of acid. The product, an ester of l-[a-(carboxy)-
benzyl or substituted benzyl]-3~-acylamido-4-acetoxy-
azetidin-2-one represented by the foregoing formula IV is
recovered from the washed solution and can be further
purified by chromatography over silica gel.
The 3~-amino-4a-acetoxyazetidin-2-ones are
prepared from the 3~-acylamido-4a-acetoxy hydrolysis product
via N-deacylation by conventional methods; for example by
reaction with phosphorus pentachloride in the presence of
pyridine to form the intermediate acylimido chloride;
reaction with methanol to form the acylimido ether; and
hydrolysis of the ether.
The 3~-acylaminoazetidin-2-ones represented by the
formula I wherein R is an acylamino group of the formula
O H
R'-C-N- are prepared via the acylation of the 3~-amino
nucleus compounds. The acylation is carried by methods
which commonly are employed in the cephalosporin and peni-
cillin art. The acylation is best carried out with an
active derivative of the carboxylic acid R'-COOH. Active
derivatives of these acids include the acyl halides such as
the acid chlorides or bromides, the acid azides, and the
mixed anhydrides formed with methyl chloroformate, ethyl
chloroformate, or iso-butyl chloroformate. Also, acylation
X-4683A -15-
,~ ,.
I
,
..
can be carried out with the free carboxylic acid with a
condensing agent such as dicyclohexylcarbodiimide as
described in U.S. Patent No. 3,218,318. The acylation is
preferably carried out via the acid halide method or via the
mixed anhydride method. The acylation via acyl halides can
be carried out in an aqueous or non-aqueous solvent in the
presence of a hydrogen halide acceptor such as sodium
bicarbonate, a tertiary amine such as triethylamine or
pyridine or an alkylene oxide such as propylene oxide or
butylene oxide. The mixed anhydride method of acylation is
carried out under anhydrous conditions in the presence of
triethylamine.
The carboxylic acids R'-C-OH employed in the
synthesis of the compounds of the formula I are readily
available either from commercial sources or via known
preparative methods.
During the preparation of the compounds of
formula I any reactive functional groups, such as the amino,
hydroxy, or carboxy groups, are protected with a suitable
blocking group. Numerous carboxy, hydroxy, and amino-
protecting groups which are employed in the penicillin and
cephalosporin art for the protection of such groups during
the chemical reactions can be employed in the preparation of
the compounds described herein. For example, the amino
group can be protected with the t-butyloxycarbonyl group or
the 2,2,2-trichloroethoxycarbonyl group; the hydroxy group
can be protected with the benzyloxycarbonyl group or a
substituted benzyloxycarbonyl group, for example, 4-
nitrobenzyloxycarbonyl or the protecting group formed by
X-4683A -16-
~ f~
rcacting the hydroxy group with methyl vinyl ether; and the
substituents which are acidic, such as the carboxy, sulfo,
or sulfamino groups, can be protected with a suitable
carboxylic acid blocking group for example an ester such as
benzyl, a substi~uted benæyl group, for example, 4-nitro-
benzyl, 4-methoxybenzyl, or 2,4,6-trimethylbenzyl, the
diphenylmethyl ester group, the trihaloethyl ester groups
for example 2,2,2-trichloroethyl, or other suitable car-
boxylic acid blocking group. These blocking groups for the
above-defined functional groups are selected from those
groups recognized as functional blocking groups which are
readily cleaved following the reaction.
The azetidin-2-one compounds represented by the
formula I wherein ~ is an aeylamino group of the formula
O H Z O
Il I ~__~ 11 11
Ra-O-C-C-CH2-CH2-O---~ ~---C-C-NH-
NH
are prepared by acylating a 3~-aminoazetidin-2-one nucleus
compound with the amino-protected and esterified 4-(3-
carboxy-3-aminopropoxy)phenylglyoxylic acid or the oxime or
protected oxime thereof. This acid and the oxime or pro-
tected oxime are prepared by the following method.
An amino protected salt of D-methionine of the
formula
O H
Il I
M-O-C-C-CH2-CH2-S-CH~
NH
Rb
X-4683~ -17-
for example, the salt wherein M is dicyclohexylammoni.um and
Rb :is as previously defined herein, is converted to the
trimethylsilyl ester and is alkylated on the sulfur atom
with an alkyl or benzyl iodide, for example methyl iodide,
and the alkylsulfonium iodide of the formula
0 H
(CH3) 3S i-0-C-C-CHz-CHz-S-CH3
NH CH3
Rb
is reacted in an inert solvent with potassium t-butoxide to
form the cyclic amino-protected D-homoserine lactone of the
formula
D-
o
The lactone is hydrolyzed with an alkali metal hydroxide to
the amino-protected D-homoserine alkali metal salt of the
formula
H H
Rb-N-C-CHz-CHz-OH
COOM~
wherein M' is sodium or potassium, and the latter is con-
verted to an acid labile ester e.g., diphenylmethyl ester.
The esterified D-homoserine is then coupled with 4-hydroxy-
phenylglyoxylic acid p-nitrobenzyl ester with a trialkyl or
triaryl phosphine, preferably triphenylphosphine, and
x-4683~ -18-
diethyl azodicarboxylate to form the amino-protected diester
of the formula
O 11 ~)
R -O-C-C-CH2-CH2-O-- ~ C-COO-CH2--~ --NO2
NH
Rb
The p-nitrobenzyl ester group is selectively de-esterified
by reduction whereby the other ester Ra, which is selected
from among the acid-labile ester groups, remains substantially
intact. For example, the p-nitrobenzyl ester group is
removed via reduction with sodium sulfide while the ester
group Ra, which can be an acid sensitive group such as the
diphenylmethyl group, remains unaffected under the reduction
conditions. The selective de-esterification product, the
phenylglyoxylic acid, is represented by the formula
O H O
Ra-O-C-C-CH%-CH2-O---~ ~---C-COOH
NH
Rb
The amino-protected and esterified phenylglyoxylic
acid is converted to an active ester which is used to
acylate a 3~-aminoazetidin-2-one nucleus compound. After
acylation the a-ketoacylamide intermediate is converted to
the biologically active oxime. For example, 4-[3-(t-
butyloxycarbamido)-3-(diphenylmethoxycarbonyl)propoxy]-
phenylglyoxylic acid is converted to the active ester formed
with l-hydroxybenzotriazole by using dicyclohexylcarbodi-
imide as condensing agent and the ester is coupled with
l-[a-(benzyloxycarbonyl)-4-benzyloxybenzyl]-3~-amino-4a-
X-4683A -19-
G:~
acetoxya~eti~in-2-one to provide l-[a-(benzyloxycarbonyl)-
4-benzyloxybenzyl]-3~-[4-(3-t-butyloxycarbamido)-3-(di-
phenylmethoxycarbonyl)propoxy]phenylglyoxylamido-4a-
acetoxyazetidin-2-one represented by the following formula.
o
O H O O H OC-CH3
Il I ~__.~ 11 11 1 .
DPM-o-C-C-CH2-CH2-O--~\ =., I I ~ -O-B~
t-BOC COO-Bz
wherein DPM = diphenylmethyl; t-BOC = t-butyloxycarbonyl;
and Bz = benzyl.
The above a-ketoamide is then converted to the
oxime derivative with hydroxylamine hydrochloride in the
presence of a weak base such as sodium bicarbonate, and the
t-BOC group, the DPM and benzyl groups removed to provide
the antibiotlc of the formula Vlas shown below.
O
H O H OC-CH3
HOOC-C-CH2-CHz-O~ c .~ ~. OH
OH CooH
Alternatively, the nocardicin side chain can be
synthesized by first forming the protected oxime of th~
4-hydroxyphenylglyoxylic acid ester and then coupling the
oxime ester with the amino-protected and esterified
D-homoserine fragment followed by selective de-esterifi-
cation of the glyoxylic acid ester. For example p-nitro-
benzyl 4-hydroxyphenylglyoxylate is reacted with hydroxyl-
X-4683A -20-
..''.' ~
I
~ f~
amino hydrochloride and the oxime is reacted with potassium
t-butoxide followed by p-methoxybenzyl bromide to form the
O-(p-methoxybenzyl)oxime. The ox:ime fragment is then
coupled by the above described method with D-3-(t-butyloxy-
carbamido)-3-(diphenylmethoxycarbonyl)propanol (an amino
protected and esterified D-homoserine) to form p-nitrobenzyl
D-4- [ 3-(t-butyloxycarbamido)-3-(diphenylmethoxycarbonyl)-
propoxy]phenylglyoxylate O-(p-methoxybenzyl)oxime repre-
sented by the formula
~OpMB
O H N
DPM-O-C--C--CHz-CH~-O--~ - C-C-O-pN8
N-H O
t-BOC
wherein DPM = diphenylmethyl, t-BOC = t-butyloxycarbonyl,
pMB = p-methoxybenzyl and pNB = p-nitrobenzyl.
The p-nitrobenzyl ester group is then removed by
chemical reduction for example with zinc and acid or by
electrolytic reduction to provide the free carboxylic acid
for acylation of the 3~-amino nucleus.
Examples of the azetidin-2-ones of formula I
wherein R is
H O
11
R''''-C - C-NH-
NH
C=O
y
X-4683A -21-
are listed in the table below with reference to the fol-
lowing structural formula.
o
O ~
Il I /O-C-CH3
I ~ VII
C=O COOH a'
y
R'''' Y a a
phenyl -N(CH3)C-NH-CH3 H H
phenyl -N(CH3)C-NH CH3 4-OH H
"
2-thienyl -N(CH3)C-NH- H H
2-furyl -u(cH3)c-NH-cH H . 3-OH
4-hydroxy- ,,
phenyl -N(CH3)C-NH-CH3 H H
phenyl -~ ~ -H 4-OH 3-Cl
phenyl -~ ~-SO2CH~ H H
o
2-thienyl -NHC-CH=CH0 4-OH 3-CH3
2-furyl -NHC-CH H H
phenyl -Nll-C-0 H 3-NH2
X-4683A -22-
,~ .
r; ~
Examples of the antibiotic compounds of formula I
wherein R is a group of the formula
O
R''''-C-C-NH-
N
Z ' '
are illustrated in the following table.
o
O H 11
11 1 ~O-C-CH3
R~ C-C-N~ VIII
0 Z'' COOH
R'1''Z'' a a'
phenyl H H H
phenyl CH3 4-OH H
phenyl CH3 H H
phenyl H 4-OH H
phenyl H 4-OH 3-Cl
20 2-thienyl CH3 H H
2-thienyl H 4-OH H
2-thienyl CH3 4-Cl H
2-thienyl CH3 4-CH3 H ~:
2-thienyl CH3 4-nitro H
2-furyl H 4-OH H
2-furylCE13 4-OH H
2-EurylC113 E1 11
2-furylC113 4-OE~ 3-Cl
2-furyl H 4-Br H
2-furyl CH3 3-CH3 4-CH3
l/acetoxy
X-4683A -23-
The a-oximino and a-methoximino substituted
azetidin-2-ones can have either the syn or anti configura-
tion and their preparation as described above can lead to
mixtures of both configurations. The preferred configura-
tion is the syn configuration.
Examples of the compounds of formula I wherein R
is the group R''-CH2-C(O)-NH- are listed in the table below
wherein reference is made to the following structural
formula.
o
0 ~1 11
l /O-C-CI-~3
IX
COOH
R'' a a'
phenyl H H
4-hydroxyphenyl 4-OH H
phenyl 4-OH H
phenyl 3-OH H
20 2-thienyl H H
2-thienyl 4-OH H
2-thienyl 4-OH 3-Cl
2-furyl H H
2-furyl 4-NH2 H
thiazol-4-yl H H
X-4~,~3~ -24-
1~' ~ ~1 '1'
isothiazol-5-yl 3-Br 11
oxazol-S-yl ~1 H
oxazol-5-yl 4-CH3 3-CH3
oxazol-5-yl 4-011 H
lH-tetrazol-l-yl H H
l}1-tetrazol-l-yl 4-NH-CH2 H
isoxazol-4-yl H H
3-methyl-4-chloro-
isoxazol-5-yl 4-OH H
3-(2-chlorophenyl)-
isoxazol-4-yl H H
3,4-dichloro-
isoxazol-5-yl 4-OH 3-Cl
3-chloro-4-methyl-
isoxazol-5-yl H H ~ :
3,5-dimethylisoxa-
zol-4-yl 4-OCH3 3-OCH3
Examples of the compounds of formula I wherein R
is the acylamino group R'''-S-CH2-C(O)-NH-are given in the
table below with reference to the following structural
formula.
o
O H 11
I l I/OC-CH3
R ' ' ' -S-CH2-C-N~ a X
COOH a
X-4683A -25-
R''' a a'
phenyl H ~1
phenyl 4-OH H
phenyl 4-Cl H
3,4-dichloro-
phenyl 4-OH 3-OCH3
3,5-dichloro-
phenyl 4-OH 3-Cl
3-methyl-4-
hydroxyphenyl 4-OH H
4-pyridyl H H
4-pyridyl H 3-OH
4-pyridyl 3-Cl H
4-pyridyl 4-Br H
4-pyridyl 3-OC2H5 4-OH
2-thiazolyl 4-OH H
1,3,4-thiadia-
zol-2-yl H H
1,3,4-thiadia-
zolyl-2-yl 4-OH 3-Cl
1,3,4-oxadia-
zol-2-yl H H
20 1,3,4-oxadia-
zol-2-yl 3-CH3 4-CH3
Examples of the compounds represented by the
formula I wherein R is a group of the formula
H O
~ 11
R ' ' ' ' -C-C-NH-
Q
are listed below with reference to the following structural
rormula .
X-4683A -26-
~I O 0 11
1111 /0-C-CHo
R~ C-C-~ /a XI
COOH a
R'''' Q a a'
phenylNH2 H H
phenylNH2 4-OH H
phenylNH2 4-OH 3-Cl
10 phenylNH2 4-Cl H
2-thienyl NH2 4-OH H
2-furylNH2 4-OH H .
2-thienyl NH2 4-OCH3 3-CH3
phenylOH 4-OH H
phenylCOOH H H
phenylSO3H 4-OH 3-OH
2-thienyl COOH H H
4-hydroxyphenyl NH2 H H
4-hydroxyphenyl COOH 4-OH 3-Cl
20 2-furyl SO3H H 2-OCH3
3-chloro-4-
hydroxyphenyl OH 4-OH H
2-thienyl OH 4-Cl H
2-thienyl -SO3H C2H5 H
4-chlorophenyl OH 4-OH 3-CH3
The 3~-acylaminoazetidin-2-ones represented by
formula I wherein R1, Ra, Rb and Z' are hydrogen, a and a'
are other than protected hydroxy, protected amino or pro- ~:
tected aminomethyl, and O is other than a protected moiety,
:30
X-4683A -27-
,
are useful antibiotics which inhibit the growth of pathogenic
microorganisms. These compounds are resistant to inactivation
by the ~-lactamases and possess activity against the gram-
negative bacteria which proliferate these enzymes, for
example, proteus, pseudomonas, enterobacter sp., serratia
and klebsiella.
The compounds are administered parenterally, for
example, subcutaneously, intramuscularly or intravenously,
preferably in the form of a pharmaceutically acceptable
non-toxic salt.
The esterified, amino-protected, oxime-protected
and hydroxy-protected azetidinones represented by the
formula I useful as intermediates in the synthesis of the
antibiotic compounds.
The azetidin-2-one antibiotics represented by the
formula I have an acidic carboxylic acid group which forms
salts with suitable bases. Pharmaceutically acceptable
salts include the alkali metal salts such as the sodium,
potassium, or lithium salt; the calcium salt; and salts
formed with pharmaceutically acceptable amines, for example,
mono- and diethanol amine, procaine, cyclohexylamine,
dicyclohexylamine, dibenzylamine, abietylamine, trimethyl-
amine, or triethylamine.
3~-Acylaminoazetidin-2-ones of the formula I
wherein Q is an amino group can form acid addition salts,
for example, the hydrochloride or hydrobromide salts.
Likewise, when a phenyl group substituent a or a' and b or
b' is amino, acid addition salts of the antibiotics can be
prepared.
:30
X-4683A -28-
Further, it will be appreciated that the 3~-amino
and 3~-acylamino which contain an a-amino substituent in the
side chain (Q = NH2) can form intramolecular salts (zwitterions)
when Rl is hydrogen.
The 3~-aminoazetidin-2-one nucleus compounds
represented by the formula I when R is an amino group are
useful intermediates in the preparation of the 3,~-acyl-
aminoazetidin-2-one antibiotics. As described previously
herein when the phenyl group of a-carboxybenzyl substituent
in the l-position of the azetidin-2-one ring is substituted
with hydroxy or amino groups, such groups are preferably
blocked with a suitable blocking group during synthesis for
example, during N-acylation or N-deacylation. These nucleus
compounds containing such blocked groups as well as esters
of the nucleus compounds are also valuable intermediates.
Examples of these nucleus compounds are listed below with
reference to the following formula.
o
O-C-CH3
~I ~c~ XII
COORl a
Rl a a
H H H
H 4-OH H
benzyl 4-benzyloxy H
benzyl 4-Cl H
DPMl H 2-Cl
pNB 4-~-pNB 3-Cl
X-4683A -29-
'
~L~16
a a
pNI3 4-benzyloxy 3-C~13
3 4-011 3-sr
pMB 2-C1130 4-CH30
H 2-Cl 6-Cl
pNB 4-NH-BZ H
benzyl 2-CH2NHBz H
l/diphenylmethyl
2/p-nitrobenzyl
3/p-methoxybenzyl
4/benzyloxycarbonyl
The carboxylic acid protecting groups represented
by Rl in the formula I are carbon esters commonly used to
temporarily protect or block the carboxylic acid function in
other ~-lactam antibiotics such as the penicillins and
cephalosporins. Examples of these ester groups include the
haloalkyl groups such as the trichloroethyl and tribromo-
ethyl groups; the benzyl and substituted benzyl groups such
as p-nitrobenzyl, p-methoxybenzyl, 3,5-dimethoxybenzyl,
2,4,6-trimethylbenzyl; diphenylmethyl (benzhydryl), 4-
methoxydiphenylmethyl; or t-butyl. Methods for the removal
of these ester groups are well known and are described in
the literature.
As previously described herein, the 3-amino or
3-acylamino substituent R in the formula I has the ~-
configuration. The 4-acetoxy substituent group is provided
in the ~-configuration by the process described herein.
X-4683A -30-
~ l~f~ ~ ~'3
The a-carboxybenzyl or a-carboxy-substituted
benzyl group substituted on the nitrogen atom of the azetidin-
2-one ring (l-position) can have either the D or L con-
figuration and the D configuration is preferred. The
process for preparing 3~-amino-4a-acetoxyazetidin-2-one
nucleus described herein provides the preferred D-configu-
ration when the phenylglycine employed in the process has
the D-configuration.
Likewise the compounds of the formula I wherein R
is the acylamino group
O H
Il I
R''''-CH-C-N-
Q
can have either the D or L configuration. The described
compounds having the D configuration are preferred.
Certain of the compounds of formula I are pre-
ferred over others. A preferred group is represented by the
formula I when R is an acylamino group of the formula
O zo
2 0Ra-O-C-CH-CH2-CH~-O ~ --C-C-NH-
NH
Rb ..
Preferred among these compounds are those wherein
Ra and Rb are hydrogen, Z is the hydroxyimino group, a is
4-hydroxy, and a' is hydrogen. An especially preferred
compound is 4a-acetoxynocardicin represented by the formula
.~()
X-46~ -31-
H O H O-C-CH3
HOOC-C-CH2-CHz-O ~ ---OH
OH COOH
A preferred intermediate useful in the preparation
of the above 4a-acetoxynocardicin is represented by the
formula
o
O H O H O-C-CH3
DPM-O-C--C--CH2 CH2 O ~ --OH XIV
o COODPM
pMB
Another preferred group of compounds of formula
I are represented when R is benzoylamino, phenylacetylamino,
or phenoxyacetylamino. These compounds are products of the
process provided herein for the synthesis of the 4-acetoxy
substituted azetidin-2-ones and can be used in the synthesis
of the 3-amino-4-acetoxyazetidin-2-one nucleus as previously
described.
The following examples are provided to further
describe the compounds and process of this invention.
Preparation 1
Preparation of 2-benzoyl-3,3-dimethyl-7-oxo-a-(4-benzyl-
oxyphenyl)-4-thia-2,6-diazabicyclo[3.2.0]heptane-6-acetic
acid benzyl ester
A slurry of 100 g. of L-cysteine in 2 1. of dry
acetone was heated at the reflux temperature for about 17
hours. After the reaction mixture wa~ allowed to co~l .o
X-4683A -32-
about 30C. the unreacted cysteine was filtered and the
reaction product, 2,2-dimethyl-4-thiazolidinecarboxylic
a~ , crystalli~e-l L~rom the ~iltrate. qhree crops o~ the
product were obtained via successive filtrations. The
combined weight of product was 83 g.
To a suspension of 16 g. (100 mM) of the product
in 300 ml. of dry acetone were added 21 ml. of propylene
oxide. Next, 11.6 ml. (100 mM) of benzoyl chloride were
added dropwise with vigorous stirring. The temperature of
the reaction mixture slowly increased from about 25C. to
about 33C. After an hour all of the thiazolidine had
dissolved and the reaction solution began to cool. When the
temperature had dropped to 30C. the reaction solution was
evaporated to yield a white solid residue. The solid was
dissolved in acetone and was diluted with hexane to crys-
tallize the product. The product, 3-benzoyl-2,2-dimethyl-
4-thiazolidinecarboxylic acid, was filtered and dried. The
dried product weighed 18.7 g.
To a solution of 48 g. (181 mmole) of 3-benzoyl-
2,2-dimethyl-4-thiazolidinecarboxylic acid, prepared by the
procedure described above in 1.5 1. of tetrahydrofuran were
added 27.8 g. (181 mmole) of l-hydroxybenzotriazole followed
by 37.4 g. (181 mmole) of dicyclohexylcarbodiimide. The
mixture was stirred for 30 minutes at room temperature. The
reaction mixture developed into a thick slurry resulting
from the precipitation of dicyclohexylurea. To the heavy
slurry was added 63 g. (181 mmole) of benzyl D-4-benzyloxy-
phenylglycinate and the reaction mixture was stirred at room
temperature for about 2 hours. The reaction mixture was
X-4683A -33-
filtered to remove the dicyclohexylurea, the filtrate
evaporated under reduced pressure, and the residue was
dissolved in ethyl acetate. The solution was washed suc-
cessively with 5% hydrochloric ac:id, water, aqueous sodium
bicarbonate, and finally with water. The washed solution
was dried, treated with carbon, and evaporated to dryness
under reduced pressure to yield 107 g. (99% yield) of N-
[benzyl a-(D-4-benzyloxyphenyl)acetate]-3-benzoyl-2,2-
dimethyl-4-thiazolidinecarboxamide.
The above thiazolidinecarboxamide ester (107 g~;
0.18 mole) was dissolved in 4 1. of benzene and 175 g. (0 72
mole) of benzoyl peroxide were added to the solution. The
solution was then heated at the reflux temperature for 4.5
hours and thereafter was cooled to room temperature. The
reaction mixture was poured over a column packed with silica
gel and the column was eluted with benzene. Excess benzoyl
peroxide passed off the column initially and on further
elution with benzene, the product was collected. The eluate
was evaporated under reduced pressure to provide the pro-
duct, N-[benzyl ~-(D-4-benzyloxyphenyl)acetate]-3-benzoyl-
2,2-dimethyl-4-thiazolidinecarboxamide-5a-benzoate, as an
oil. The product was obtained crystalline from diethyl
ether.
The above product is represented by the following
structural formula
X-4683A -34-
O ~./
o-c~ o
o--c--~
o=c
HN-~H~ --OCH
1COCH2--~ ~-
NMR (T60, CDC13): 2.16 (s, 2CH3), 5.08 and 5.16 (2s, 2CH2),
5.13 (s, CH), 5.54 (d, CH), 6.54 (s,
CH) and 6.83-8.16 (m, aromatic H and NH)
delta.
A solution of 25.5 g. (35.8 mmole) of the thia-
zolidine-4-carboxamide 5a-benzoate in 1 1. of dry methylene
chloride was cooled to a temperature of 0C. and hydrogen
chloride was bubbled through the cold solution for about 2
hours. After this time, a thin layer chromatogram developed
with benzene:ethyl acetate, 7:3, v:v, showed complete
reaction. The methylene chloride was evaporated under
reduced pressure providing the product as a foam. The foam
was dissolved in ethyl acetate and the solution was washed
with a dilute aqueous solution of sodium bicarbonate and
with brine, and after washing was dried over magensium
sulfate, treated with carbon, and evaporated to dryness
under reduced pressure. The product, N-[benzyl a-~D-4-
benzyloxyphenyl)acetate]-3-benzoyl-2,2-dimethyl-Sa-chloro-
4-thiazolidinecarboxamide, represented by the following
formula was obtained as 22.4 g. of white foam.
X-4683A -35-
~Lr~-6
o ~.,/
C-
~
-- ---------- .. C I
o=c
HN-~CH--~ ~--OCH2-~
==~
OOCH2--~ ~-
NMR (T60, CDC13): 2.12 (s, CH3), 2.26 (s, CH3), 5.08 and
5.16 (2s, 2CH2) 5.16 (s, CH), 5.44 ~d,
CH), 5.83 (s, CH) and 6.08-7.5 (m, 20H,
aromatic H and NH) delta.
To a solution of 22.4 g. (35.8 mmole) of the 5a-
chloro-4-thiazolidinecarboxamide in 800 ml. of methylene
chloride and 200 ml. of dimethylformamide were added 1.72 g.
of sodium hydride (50 percent in oil, 35.8 mmole). The
reaction mixture was stirred at room temperature for ap-
proximately 50 minutes after which time TLC (benzene:ethyl
acetate, 7:3) showed the reaction was complete. Two milli-
liters of acetic acid were added to the reaction mixture todestroy any excess sodium hydride, and the reaction mixture
was poured into 5 percent hydrochloric acid. The organic
phase was separated and was washed with 5 percent hydro-
chloric acid and with water before drying over magnesium
sulfate. The dried extract was treated with carbon, fil-
tered, and evaporated to dryness under reduced pressure.
The residue was dissolved in about 30 ml. ethyl acetate. On
standing, 10.1 g. (crop 1) of product, 2-benzoyl-3,3-
dimethyl-7-oxo-a-(4-benzyloxyphenyl)-4-thia-2,6-diaza-
X-4683A -36-
`:
~&~:~$
bicyclo[3.2.0]heptane-6-acetic acid benzyl ester, crys-
tallized. The crystals were filtered and the filtrate was
treated with petroleum ether to the cloud point. On
standing and with agitation, 5.5 g. (crop 2) of additional
product crystallized. The second crop material was filtered
and the filtrate was evaporated to dryness to yield further
product as a foam. The foam was treated with a mixture of
ethyl acetate and ethyl alcohol which afforded 2.2 g. of
additional crystalline product (crop 3).
An analysis of the nuclear magnetic resonance
spectrum (T-60) of the above crops demonstrated that crop 1
was the D isomer of the cyclized product represented by the
formula shown below, crop 2 was the L isomer, while crop 3
was a mixture of the two isomers.
O ~/
-C-~
COOCH2-~ ~-
NMR (T60, CDC13): 1.70 (s, CH3), 1.91 (s, CH3), 5.08 and
5.16 (2s, 2CH2), 5.45 (s, CH), 5.52 (m,
2CH), and 6.83-7.67 (m, l9H, aromatic H)
delta.
The above thiazolidine azetidinone was also pre-
pared from the 5~-chlorothiazolidine carboxamide and the
base diazabicyclo[5.4.0]undec-5-ene(DBU) in the following
manner.
X-4683A -37-
~lr~6,~;~
A solution of 1.01 g. (1.6 mmole) of ~he 5~-
chloro-4-thiazolidinecarboxamide having the D-configuration
in 50 ml. of methylene chloride was cooled to a temperature
of about 0C. To the cold solution was added 0.243 g. (1.~
mmole) of DBU. The reaction mixture was stirred for 2 hours
at 0C. and then was washed with 5 percent hydrochloric acid
and with brine and was then dried over magnesium sulfate.
The solution was evaporated to yield a crude reaction
product mixture. The mixture was crystallized from benzene/
petroleum ether and the crystals filtered. The infrared
spectrum of the product showed an absorption peak at 1775 cm 1
for the ~-lactam carbonyl, while the nuclear magnetic
resonance spectrum and circular dichroism showed the
material to be optically pure.
Example 1
Preparation of l-[-(benzyloxycarbonyl)-4-benzyloxybenzyl]-
3~-benzamido-4-acetoxyazetidin-2-one.
A slurry of 300 mg (0.505 mmole) 2-benzoyl-3,3-
dimethyl-7-oxo--(4-benzyloxyphenyl)-4-thia-2,6-diazabi-
cyclo[3.2.0]heptane-6-acetic acid benzyl ester in 25 ml. of
acetic acid was stirred and treated with 240 mg. (0.75
mmole) of mercuric acetate. The reaction mixture was heated
on the steam bath for 10 minutes, and an additional 100 mg.
of mercuric acetate was added and heating was continued for
5 minutes. The reaction mixture was filtered to remove the
insoluble mercurous acetate and was then evaporated to
dryness. The residue was dissolved in ethyl acetate and the
solution was washed with a dilute aqueous solution of sodium
bicarbonate and with brine, was dried and then evaporated to
~:-4683A -38-
$
dryness to yield l-[a-(benzyloxycarbonyl)-4-benzyloxy-
benzyl]-3-(N-propenylbenzamido)-4-a-acetoxyazetidinone~2
represented by the following formula.
~CH2 OAc
~__~ Il /
H
COOCH2--~
\.= . .
The product was dissolved in 50 ml. of tetra-
hydrofuran and 5 ml. of 5 percent hydrochloric acid were
added. After stirring for 15 minutes at room temperature
the solution was evaporated to dryness under reduced pres-
sure and the residue was dissolved in ethyl acetate. The
ethyl acetate solution was washed with a dilute aqueous
solution of sodium bicarbonate, with water, and was dried
and evaporated to dryness. The residue was chromatographed
on a preparative silica gel thin layer plate using benzene:
ethyl acetate, 7:3, v:v, for development to obtain 144 mg.
of the hydrolysis product 1-[a-(benzyloxycarbonyl)-4-
benzyloxybenzyl]-3~-benzamido 4-a-acetoxyazetidinone-2.
Example 2
By following the procedures described by Example 1
l-[a-(benzyloxycarbonyl)-4-benzyloxybenzyl]-3~-phenoxy-
acetamido-4a-acetoxyazetidin-2-one is prepared with the
benzyl ester of 2-phenoxyacetyl-3,3-dimethyl-7-oxo-a-
(4-benzyloxyphenyl)-4-thia-2,6-diazabicyclo[3.2.0]heptane-
6-acetic acid.
X-4683A -39-
Examplc 3
~rc~ r(ltLon Or 1-1a-(bcnzyloxycarbonyl)-4-bcnzyloxybenzyl]-
3~-amino-4a-acetoxyazetidin-2-one.
To a solution of 250 mg. (0.432 mmole) of 1-
[a-(benzyloxycarbonyl)-4-benzyloxybenzyl]-3~-benzamido-
4-a acetoxyazetidinone 2 prepared as described in the
foregoing Example 1 in 50 ml. of dry benzene were added
132 mg. (0.64 mmole) of phosphorus pentachloride and 50 mg.
(0.64 mmole) of pyridine. The mixture was heated to a
temperature of about 65C. for 2 hours with stirring. A
thin layer chromatogram was then run on the reaction mixture
and demonstrated the conversion of the starting material to
the corresponding imido chloride.
The reaction mixture was evaporated under reduced
pressure and 50 ml. of dry methyl alcohol were added to the
residue. The methyl alcohol solution was stirred at room
temperature for about 30 minutes. TLC of the solution
showed a new spot.
The solution was evaporated under reduced pressure
and 20 ml. of water and 20 ml. of tetrahydrofuran were added
to the residue and the resultant solution was stirred at
room temperature for about 20 minutes. The tetrahydrofuran
was evaporated from the solution and the aqueous concentrate
was slurried with ethyl acetate. The pH of the slurry was
adjusted to pH 7 and the ethyl acetate layer was separated.
The ethyl acetate layer was dried and then evaporated under
reduced pressure to afford 210 mg. of a reduction product
mixture. The nuclear magnetic resonance spectrum (T-60) of
the product mixture showed it contained approximately 60
.-46~3A _40_
.: ,
. ~ w
~. :
'
percent of the desired N-deacylation product, l-[a-(benzyl-
oxycarbonyl)-4-benzyloxybenzyl]-3-~-amino-4-a-acetoxy-
azetidinone-2 represented by the following structural
formula.
H~N oAc
o/ I c ~ ,. OCH2~
COOCH2~
The crude product was purified via preparative
thin layer chromatography on silica gel thick layer plates
to yield 62 mg. of the product and 60 mg. of recovered
starting material.
Example 4
Preparation of l-[a-(carboxy)-4-hydroxybenzyl]-3~-(D-
mandelamido)-4a-acetoxyazetidin-2-one.
To a solution of l-[a-(p-nitrobenzyloxycarbonyl)-
4-benzyloxybenzyl]-3!3-amino-4a-acetoxyazetidin-2-one in
tetrahydrofuran maintained at 0-5C. are added pyridine and
O-formyl mandeloyl chloride. The mixture is stirred in the
cold for about 2 hours and is then evaporated. Water is
added to the residue and the acylation product is extracted
with ethyl acetate. The extract is washed with water,
dilute acid, and again with water and is dried and then
evaporated to yield the esterified N-acylation product,
l-[a-(p-nitrobenzyloxycarbonyl)-4-benzyloxybenzyl]-3~-
(O-formylmandelamido)-4a-acetoxyazetidin-2-one. The ester
is de-esterified with zinc and acetic acid during which the
4-benzyloxy group is removed to provide the title compound.
X-4683A -41-
~131 C~
Example 5
Preparation of l-[a-(carboxy)-4-hydroxybenzyl]-3~-[2-[4-
(3-carboxy-3-aminopropoxy)phenyl]-2-hydroximinoacetamido]-
4a-acetoxyazetidin-2-one.
To a solution of 32 mg. of l-[a-(benzyloxycar-
bonyl)-4-benzyloxybenzyl]-3~-amino-4a-acetoxyazetidin-
2-one in about 5 ml. of dry methylene chloride were added
36 mg. of 4-(3-diphenylmethoxycarbonyl-3-t-butyloxycar-
bamidopropoxy)phenylglyoxylic acid. The mixture was stirred
at room temperature under nitrogen and 14 mg. of dicyclo-
hexylcarbodiimide were added. The solution immediately
turned a dark yellow. After 10 minutes the solution became
light yellow and dicyclohexylurea precipitated. A thin
layer chromatogram of the mixture run on silica gel plates
with benzene:ethyl acetate, 7:3, v:v, showed complete
reaction. The reaction mixture was filtered and the fil-
trate was evaporated to dryness.
The acylation product was purified on preparative
thick layer plates (silica gel) to yield 41 mg. of the
purified intermediate product.
The product was reacted with hydroxylamine hydro-
chloride in aqueous tetrahydrofuran containing pyridine to
form the hydroximino derivative. The oxime was reacted with
trifluoroacetic acid in anisole to cleave both the t-BOC
amino-protecting group and the diphenylmethyl ester group of
the side chain. After treatment with the acid, the reaction
mixture was evaporated to dryness and the residue was
vigorously triturated with diethyl ether. The insoluble
product, l-[-(benzyloxycarbonyl)-4-benzyloxybenzyl]-3~-
X-4683A -42-
[2-[4-(3-carboxy-3-aminopropoxy)phenyl]-2-hydroxyimino-
acetamido]-4a-acetoxyazetidin-2-one trifluoroacetate salt,
was filtered and dried.
The salt was dissolved in dry methylene chloride
and was treated with a solution of excess aluminum chloride
and anisole in nitromethane to effect the removal of both
benzyl ester groups. The reaction mixture was quenched with
water and the pH adjusted to about 8.0 with sodium bicar-
bonate. The solution was washed with ethyl acetate and was
*
then chromatographed over Sephadex. The title compound was
obtained as the disodium salt by evaporation of the eluate.
The course of the debenzylation reaction is
followed by thin layer chromatography over silica gel using
acetic acid:acetone, 4:1, v:v, for development.
Example 6
Preparation of l-[a-(diphenylmethoxycarbonyl)-4-hydroxy-
benzyl~-3!3-amino-4a-acetoxyazetidin-2-one.
A solution of 4 g. of 2-benzoyl-3,3-dimethyl-
7-oxo-a-(4-hydroxyphenyl)-4-thia-2,6-diazabicyclo[3.2.0]-
heptane-6-acetic acid and 5 g. of diF~henyldiazomethane in
500 ml. of dry tetrahydrofuran was stirred at room tem-
perature for about 18 hours. The reaction mixture was
evaporated under reduced pressure to dryness and the residue
was dissolved in ethyl acetate. The diphenylmethyl ester
product, 2-benzoyl-3,3-dimethyl-7-oxo-a-(4-hydroxyphenyl)-
4-thia-2,6-diazabyciclo[3.2.0]heptane-6-acetic acid diphenyl-
methyl ester, crystallized from solution. 3.5 Grams of the
diphenylmethyl ester were obtained.
X-4683A -43-
*Trademark of Pharmacia Fine Chemicals, Inc. for~a cross-
linked dextran (polysaccharide) gel in bead form,
employed in chromatography.
To a suspension of 1 g. of the diphenylmethyl
ester in approximately 100 ml. of acetic acid were added
with stirring 1 g. of mercuric acetate. The reaction
mixture was heated on the steam bath for approximately 10
minutes with continued stirring. The reaction mixture was
filtered to remove the insolubles and was evaporated under
reduced pressure to dryness. The reaction product mixture
obtained as a residue was dissolved in ethyl acetate and the
solution was washed with a dilute aqueous solution of sodium
bicarbonate and with brine and was then dried and evaporated
to dryness to yield 1.1 g. of l-[a-(diphenylmethoxycar-
bonyl)-4-hydroxybenæyl]-3-(N-propenylbenzamido)-4a-acetoxy-
azetidinone-2.
The propenyi 4a-acetoxyazetidin-2-one diphenyl-
methyl ester, 650 mg. was dissolved in 50 ml. of water and
50 ml. of tetrahydrofuran and 650 mg. of mercuric acetate
were added. The reaction mixture was stirred at about room
temperature for approximately 1 hour. The reaction mixture
was evaporated to dryness under reduced pressure and the
residue containing the hydrolysis product was dissolved in
ethyl acetate. The ethyl acetate solution was washed with a
dilute aqueous solution of sodium bicarbonate, water, and
was then dried and evaporated to dryness. The nuclear
magnetic resonance spectrum (T60) showed that complete
hydrolysis of the N-propenyl group had taken place. The
product, 600 mg., 1-[a-(diphenylmethoxycarbonyl)-4-hydroxy-
benzyll-3~-benzamido-4a-acetoxyazetidinone-2 was used
without further purification as described hereinafter.
X-4683A -44-
61-~3
To a solution of 510 mg. of the 4a-acetoxyazeti-
dinone-2-diphenylmethyl ester prepared as described above in
40 ml. of dry tetrahydrofuran were added 2 ml. of dihydro-
pyran and a small catalytic amount of p-toluenesulfonic
acid. The reaction mixture was stirred at about room
temperature for approximately 17 hours. A silica gel thin
layer chromatogram showed the presence of starting material
and the desired product, the tetrahydropyranyl ether formed
with the 4-hydroxybenzyl group. With continued stirring
approximately 0.5 g. of sodium carbonate were added to the
reaction mixture. After stirring for 15 minutes the solvent
tetrahydrofuran was evaporated and the residue was dissolved
in ethyl acetate. The ethyl acetate solution was washed
with sodium bicarbonate and with water and was then dried
and evaporated to dryness. The starting material and
product contained in the residue were separated on a silica
gel chromatographic plate employing 7:3, toluene:ethyl
acetate, v:v, for elution. There were obtained 280 mg. of
the product, 1-[a-(diphenylmethoxycarbonyl)-4-tetrahydro-
pyranyloxybenzyl]-3~-benzamido-4a-acetoxyazetidinone-2 and
178 mg. of recovered starting material.
To a solution of 78.5 mg. (0.38 mmole) of phos-
phorus pentachloride and 29.9 mg. (0.38 mmole) of pyridine
in approximately 20 ml. of dry methylene chloride were added
with stirring at room temperature under nitrogen 168 mg.
(0.26 mmole) of the 4a-acetoxyazetidin-2-one tetrahydro-
pyranyl ether diphenylmethyl ester. The reaction mixture
was stirred at room temperature under nitrogen and peri-
X-4683A -45-
odically an aliquot was withdrawn from the reaction mixture
and chromatographed on thin layer chromatography plates
using benzene:ethyl acetate, 7:3 for elution. After
approximately 1 hour the TLC showed only a trace of starting
material and a new spot corresponding to the imino chloride.
The methylene chloride was evaporated and 30 ml. of dry
methyl alcohol were added to the concentrate. The methyl
alcohol solution was stirred for approximately 2 hours at
about room temperature. Thereafter, the methanol was
evaporated and 20 ml. of water and 20 ml. of tetrahydrofuran
were added to the residue. The aqueous solution was stirred
for approximately 1 hour at about room temperature and
thereafter the solution was evaporated to remove the tetra-
hydrofuran. The aqueous concentrate was extracted with
ethyl acetate and the extract was dried and evaporated to
dryness. The product obtained as a crude residue was
purified via chromatography over silica gel to yield 81 mg.
of l-[a-(diphenylmethoxycarbonyl)-4-hydroxybenzyl]-3~-
amino-4~-acetoxyazetidin-2-one.
Example 7
Preparation of 4a-acetoxynocardicin.
To a solution of 300 mg. (0.65 mmole) of the
4~-acetoxyazetidin-2-one nucleus diphenylmethyl ester pre-
pared as described in the preceding example and 435 mg.
(0.65 mmole) of 4-[3-(diphenylmethoxycarbonyl)-3-(t-
butyloxycarbamido)propoxy]phenylglyoxylic acid 0-(4-
methoxybenzyl)oxime in approximately 25 ml. of dry methylene
chloride maintained at about room temperature under nitrogen
were added 135 mg. (0.65 mmole) of dicyclohexylcarbodiimide.
X-4683A -46-
~16~
The reaction mixture was stirred at approximately room
temperature under nitrogen for about 2 hours. The reaction
mixture was filtered and the filtrate was evaporated to
dryness. The residue was chromatographed on a preparative
silica gel chromatographic plate employing toluene:ethyl
acetate, 1:1, v:v for elution to obtain 391 mg. of the
acylated product, the t-butyloxycarbonyl protected amino,
bis-dibenzhydryl ester protected and 4-methoxybenzyl protected
oxime derivative of 4a-acetoxynocardicin represented by the
formula below. The NMR of the product (T60) showed a trace
impurity. The product was further purified in the same
chromatographic system affording 332 mg. of the purified
product.
o
O H O O-C-CH3
PM13 COODPM
The protected bis-diphenylmethylester of 4a
acetoxynocardicin of the above formula was treated with
12 ml. of trifluoroacetic acid containing 24 drops of anisole
at approximately room temperature for 3 minutes. The
reaction mixture was evaporated to dryness and the residue
was vigorously triturated with diethyl ether to yield 157 mg.
of 4a-acetoxynocardicin trifluoroacetate salt.
NMR: DMSO(D6), D2O (Trimethylsilane reference, 1.95
(s, -C(O)CI13), 2.31 (t, CH2), 4.15 (t, and d,
-CH-CH2), 4.74 (d, -CH-), 5.20 (s, -CH-), 5.88
(d, -CH-) and 6.66-7.52 (aromatic H) delta.
X-4683A -47-
