Note: Descriptions are shown in the official language in which they were submitted.
The present in~ention relates to a steroselective
total synthesis of certain novel substituted a2 '3-1, 4-
morpholine-2-carboxylic acids possessing a fused ~-lactam ring
in the 1,6-position and carrying a substituent cis to car~on
5 in the 7-position of the fused ring system represented by
the general formula
H H 5
~C~::25-Z
C02H
wherein X is amino, azido or acylamido and Z represents
optionally substituted Cl-C6 alkyl, aryl, aralkyl or
heterocyclic. When X is acylamino, these acids (and their
pharmaceutically acceptable salts and physiologically
hydrolyzed esters) are potent antibacterial agents.
Also included in this invention are various
novel intermediates useful in preparing the active ~-
lactam derivatives desribed above and various processes
for the production of the intermediates and active com-
pounds.
The compounds having the above general form-
ula represent a new family of ~-lactam antibiotics. They
can be considered nuclear analogs of cephalosporins in
which the sulphur atom of the dihydrothiazine ring is
replaced by an oxygen atom and shifted from position
5 to position 4 of the ~-lactam ring system as numbered
in the formula above.
-1-
~1~3~3~ '
Sheehan has used the term 0-cepham for the structure
7 ~ 0
o
0-Cepham
[J.C. Sheehan and M. Dadic, J. Heterocyclic Chem., 5,
770 (1968)] and we propose the use of the term 0-2-
isocepham for the basic system having the formula
7 6 ~
~ 0 2
; ~ .
The numerical prefix indicates the position of the
hetero-atom.
There is thus provided by the present invention
the novel 0-2-isocephem carboxylic acid compounds having
10 the formula
H H
~ 7
R-NH l l 0
~ N ~ CH2-S-Z
C02H
II
wherein R is an acyl group and Z represents optionally
substituted Cl-C6 alkyl, aryl, aralkyl or heterocyclic,
and easily cleavable esters and pharmaceutically
acceptable salts of said acids and esters.
- 2 -
3~
The acyl group R can be chosen from a wide variety
of organic acyl radicals which yield products of
: improved properties and is preferably an acyl radical
which is contained in a naturally occurring or bio-
synthetically, semi-synthetically or totally-syntheti- -
cally produced pharmacologically active N-acyl derivative
of 6-arninopenicillanic acid or 7-aminocephalosporanic
acid. Examples of suitable acyl groups which can be used
have been defined in our related Belgian Patent 837,265,
but it should be noted that this is not intended to be an
exhaustive list of all the possible acyl groups which may
be used.
A preferred class of acyl groups are those of
the forrnula
O
Ar'-CH-C
. .
: wherein Ar' is a radical of the formula
Im Rm
Rn~
R
(~ o
i~8~3~
in which ~m, ~n ~nd R are alike or different and each
is hydrogen, hydroxy, (lower)alkyl, cyano, (l~wer)alkoxy,
chloro, bromo, iodo, fluoro, trifluoromethyl, nitro,
amino, (lower)alkylamino, di(lower)alkylamino, (lower)-
alkanoyl, tlower)alkanoyloxy such as p-acetoxy or phenyl
and Y is amino or a group cbtained by reacting the amino
group with acetaldehyde, formaldehyde or acetone, fluoro,
chloro, bromo, iodo, hydroxy, (lower)alkanoyloxy, carboxy,
guanidino, 3-guanyl-1-ureido, 3-(2-furoyl)ureido, 3-benzoyl-
ureido, sulfo, sulfoamino, ureido, thioureido, (lower)-
alkoxy, cyano, cyanamino or indanyloxycarbonyl. Parti-
cularly preferred Ar radicals are phenyl, p-hydroxyphenyl,
4-hydroxy-3,5-dichlorophenyl, 3-chloro-4-hydroxyphenyl,
o-, m- or p- aminomethylphenyl, 2-thienyl, 3-thienyl,
l-cyclohexenyl and 1,4-cyclohexadienyl. Particularly
preferred Y groups are amino, hydroxy and carboxy.
Set forth below are formulae of the most preferred acyl
groups of this class:
~ IN-CO- ; ~o ~ CN-CO-
,
- 4 -
(~-? ` O
813S
Cl
~ Cl
H~/ \~--CN-CO- HO
C;S~CH^CO- ;
N-CO- H-CO-
NH2
NH2 ~IH2
~CH-CO- ;
1H
E3--CH-CO- ICH_CO_
OH
~3~ H-CO- ; ~:H-CO-
OH OH
.
~ IH-co_ ~CH-co-
.
5 --
813S
CH-CO- ~CIH-CO-
S I ; COOH
COOH
~-CO- ~ 3-CO-
COOH
,~
Of most interest are the acyl groups of the above class
where the acid ArCH(X)COOH is of the D-series.
Other particularly preferred acyl groups for
the compounds of formula I are
OCH3
N ' C-CH2CO- ; ~ CO-
-: OCH3
N = N
~ N-CH CO- ; ~
N = C N ~ S~H2CO-;
- U
: ~ C~3
where U and V are alike or different and each is
hydrogen, chloro or fluoro;
- 6
`:
:
llQ8135
~(:H2C{l- CH2CO- ' -
~H2CO- ,~--CH2C-
~C-CH2CO~ CH-CO-
I
O _ C -NH-C= O
16HS
OCH2CO- . ~SCH2CO- '
9~2NH2
2CO- , Cl-C7 alkyl-CO-
~CO- ' ~CH2CO-
OC2~15
~3_ . ~C~12CO~
CH2CO , N~,N
Cl-C7 slkoxy-CO-, O~CO- ~ I
~CH2C-IIH-CH2CO-, I
NH ~OC~2NHCO- '
OH
O--C-- C--H ~h
C \~ CH2CO~
~N~/--CH2CO-
; ' ;
~ , ~CO-
'
CO- .~ SCH2CO- ,
~_<CH2NH2 CH2NH2
~C~12CO- 9nd~--CH2CO-
,
--8-- . .
l~J8~135
Substituent Z in formulae I and II above may
be Cl-C6 alkyl, aryl, aralkyl or heterocyclic, any of
said groups being optionally substituted by one or
more substituerts. The alkyl group may be a monova-
lent saturated aliphatic hydrocarbon radical having
from 1 to 6 carbon atoms and a straight or branched
chain, e.g., methyl, ethyl, propyl, isopropyl, butyl,
isobutyl, t-butyl, amyl or hexyl. The aryl group
may be a mono-, bi- or polycyclic aromatic hydrocarkon
radical, e.g., phenyl, l-naphthyl, 2-naphthyl or 2-
phenanthryl. The term aralkyl as used herein includes
monovalent aryl-substituted aliphatic hydrocarbon
radicals o~ the formula aryl-(ALK)m~ in which aryl
is as defined above, m is an integer of 1 to 4 and
ALK represents a straight or branched chain alkylene
radical, e.g., -(ALK)m- may be methylene, ethylene,
propylene, butylene, l-methylpropylene, 2-ethylethylene
and the like. Eeterocyclic Z substituents may be
heteromonocyclic or heterobicyclic residues of
aromatic character as well as appropriate partially
or wholly saturated residues.
A preferred group of compounds of formulae
I and II are those in which Z is Cl-C6 alkyl, aryl
selected from phenyl or naphthyl, aralkyl of the
formula aryl-(ALK)m- in which aryl is phenyl or
naphthyl, m is an integer of 1 to 4 and ALK represents
8135
a straight or branehed ehain alkylene radieal, or
heteroeyelie seleeted from a S- or 6- membered
heteroeyclic ring eontaining 1 to 4 atoms seleeted
from N, O or S, said alkyl radi~al being optionally
substituted by one or more substituents seleeted from
hydroxy, halo, amino, nitro, di(Cl-C4 alkyl) amino,
earboxy, sulfo or eyano and said aryl, aralkyl or
heteroeyelic radicals being optionally substituted
by one or more substituents selected from halo,
Cl-C4 alkyl, Cl-C4 alkoxy, eyano, earboxyl, amino,
nitro, C3-C4 eycloalkyl, C2-C4 alkenyl, trifluoromethyl,
hydroxy, hydroxymethyl~ Cl-C4 alkylthio, Cl-C4
~ alkylamino, di(Cl-C4 alkyl) amino, mercapto, phenyl,
:: benzyl, alkoxyalkyl of up to 4 earbons or -(CH2)nCOOH
. in which n is an integer of 1 to 4. Examples of suit-
able heterocyelic radicals include thienyl, furyl,
~ pyrazolyl, imidazolyl, isoimidazolyl, triazolyl,
i tetrazolyl, thiazolyl, thiadiazolyl, oxazolyl, oxadiazolyl,
isothiazolyl, isoxazolyl, pyridyl, pyridazinyl,
pyrazinyl, pyrimidinyl and triazinyl.
A most preferred group of compounds of
formulae I and II are those in which Z is an optionally
substituted 5- or 6- membered heterocyclic ring eontaining
1 to 4 atoms seleeted from N, O and S, the substituents
being preferably those mentioned above. Ecpecially
preferred heterocyelie rings are optionally substituted
'
-- 10 --
8 3 3 5
triazole, thiadiazole, oxadiazole or tetrazole
radicals. Within this group the most preferred
Z substituents are 1,2,3-triazolyl-5-yl, 2-methyl-
1,3,4-thiadiazol-5-yl, 2-methyl-1,3,4-oxadiazol-
5-yl, l-N-methyltetrazolyl, l-carboxymethyltetra-
zol-5-yl and 1-carboxyethyltetrazol-5-yl.
The term "(lower)alkyl" as used herein
means both straight and branched chain aliphatic hydro-
carbon radicals having from one to ten carbon atoms
such as methyl, ethyl, proyl, isopropyl, butyl, isobutyl,
t-butyl, amyl, hexyl, 2-ethylhexyl, heptyl, decyl, etc.
Similarly, where the term "(lower)" is used as part
of the description of another group, e.g. "(lower)-
alkoxy", it refers to the alkyl portion of such group
which is therefore described above in connection with
"(lower)alkyl".
The pharmaceutically acceptable salts re-
ferred to above include the nontoxic carboxylic acid
salts, e.g. nontoxic metallic salts such as sodium,
potassium, calcium and aluminum, the ammonium salt and
salts with nontoxic amines, e.g. trialkylamines, procaine,
dibenzylamine, N-benzyl-~-phenethylamine, l-ephenamine,
N,N'-dibenzylethylenediamine, N-alkylpiperidine and
other amines which have been used to form salts of
penicillins and cephalosporins. When a basic group
is present, as when it occurs in the 7-acyl group,
the present invention also includes the pharmaceutically
acceptable`acid addition salts, e.g. salts with mineral
-- 11 --
~8~S
acids such as ~ydrochloric, hydrobromic, hydroiodic,
phosphoric, sulfuric and salts with organic acids such
as maleic, acetic, citric, oxalic, succinic, benzoic,
tartaric, fumaric, mandelic, as-orbic ar,d malic. ~he
term "pharmaceutically acceptable salts" is also
meant to include nontoxic acid addition salts of
the easily cleavable esters referred to above. The
compounds which contain a basic group in radical
R may also be present in the form of an internal
salt, i.e. in the form of the zwitterion.
The easily cleavable esters referred to
above include ester groups which are removable by
methods, e.g. chemical or enzymatic hydrolysis, txeat-
ment with chemical reducing agents under mild condi-
tions, irradiation with ultraviolet light or catalytic
hydrogenation, which do not result in any appreciable
destruction of the remaining portion of the molecule.
Examples of suitable esters include those disclosed
in U.S. Patents 3,284,451 and 3,249,622 and U.K.
Patents 1,229,453 and 1,073,530. Esters which have
been used previously in penicillin and cephalosporin
chemistry include for example benzhydryl, p-nitrobenzyl,
benzyl, trichloroethyl, silyl such as trimethylsilyl,
phenacyl, p-methoxybenzyl, acetonyl, phthalidyl, indanyl
and (lower)alkyl such as methyl, ethyl and t-butyl.
Particularly preferred easily cleavable esters are
those which are hydrolyzed under physiological condi-
tions such as pivaloyloxymethyl, acetoxymethyl, phthalidyl,
indanyl and methoxymethyl.
- 12 -
1~8~
As the 0-2-isocephem compounds of the
present invention may possess one or more asymmetric
carbon atoms, the invention includes all of the pos-
sible enantiomeric and diastereomeric forms of th~ com-
pounds of the general formula II shown above. Resulting
mixtures of isomers can be sepaxated into the
individual isomers according to methods which are
known per se, e.g. fractional crystallization, adsorp-
tion chromatography or other suitable separation
processes. Resulting racemates can be separated into
the antipodes in the usual manner after introduction
of suitable salt-forming groupings, e.g~ by forming
a mixture of diastereoisomeric salts with optionally
active salt-forming agents, separating the mixture
into diastereoisomeric salts, and converting the
separated salts into the free compounds, or by frac-
tional crystallization from optically active solvents.
It will be appreciated that certain of the
compounds of this invention exist in various states
of solvation and the anhydrous as well as solvated
forms are within the scope of the invention.
The free acid compounds of general formula
II and general formula I wherein R is acylamido
and physiologically hydrolyzed esters thereof
together with the pharmaceutically acceptable
salts of such free acids and esters are useful
as antibacterial agents. The remaining acids,
esters and salts of formula I are valuable inter-
- 13 -
mediates which can be converted into the pharma-
cologically active compounds of formula II as
by the prscesses described below.
Preferred compounds of formula II are
those in which R is an organic acyl group. Use of
the acyl groups mentioned in Belgain patent 837,265 as
well as those mentioned above results in active end-
products having advantageous pharmacological properties.
A preferred embodiment of the present
invention consists of the compounds of formula
H H
R-NH ~ o
'.; I I I
O I CH2-S-Z
. C02H
II
in which R is an organic acyl group as defined above
and Z represents a 5- or 6-membered heterocyclic
ring containing N, O or S, said heterocyclic ring
being optionally substituted by one or more
,
,i'
,
- 14 -
~ 8135
substituents selected from halo, Cl-C4 alkyl,
Cl-C4 alkoxy, cyano, carboxyl, amino, nitro,
C3-C4 cycloalkyl, C2-C4 alkenyl, trifluoromethyl,
hydroxy, hyd.roxymethyl, Cl-C4 alkylthio, Cl-C4 alkyl-
amino, di (Cl-C4 alkyl) amino, mercapto, phenyl,
benzyl, alkoxyalkyl of up to 4 carbons or ~CH2)nCOOH
in which n is an integer of 1 to 4, and easily cleavable
esters and pharmaceutically acceptable salts thereof.
A most preferred subclass within the above
compounds of Formula II consists of the D-isomers of
the acids in which R is ~-amino-~-(p-hydroxyphenyl)acetyl,
~-amino-~-(3-chloro-4-hydroxyphenyl)acetyl, ~-amino-~-
(3,5-dichloro-4-hydroxyphenyl)acetyl, -amino-~-(2-
thienyl)acetyl, ~-amino-~-(3-thienyl)acetyl, ~-amino-a-
(1-cyclohexenyl)acetyl, ~-amino-~-(1,4-cyclohexadienyl)-
acetyl, ~-hydroxyacetyl, ~-hydroxy-~-(2-thienyl)acetyl,
a-hydroxy-~-(3-thienyl)acetyl, ~-hydroxy-~-(l-cyclohexenyl)-
acetyl or a ~-hydroxy-~-(1,4-cyclohexadienyl)acetyl, or
pharmaceutically acceptable salts thereof.
-- 15 --
11~8~35
Another preferred embodiment of the present
invention consists of the compounds of formula
H H
R-NH ~ o
~ ' I I I . ~.
~ ~\ ';
O ¦ CH2-S-Z
Co2H
wherein R is an organic acyl group as defined above and
Z is a triazole, thiadiazole, oxadiazole or tetrazole
radical, said radical being optionally substituted by
one or more substituents selected from halo, Cl-C4
alkyl, Cl-C4 alkoxy, cyano, carboxyl, amino, nitro,
C3-C4 cycloalkyl, C2~C4 alkenyl, trifluoromethyl, hydroxy,
hydroxymethyl, Cl-C4 alkylthio, Cl-C4 alkylamino, di
(Cl-C4 alkyl)amino, phenyl, benzyl, mercapto, alkoxyalkyl
of up to 4 carbons or -(CH2)nCOOH in which n is an
integer of 1 to 4, and easily cleavable esters and
pharmaceutically acceptable salts thereof.
A most preferred embodiment of the present
invention consists of the compounds of formula II in
which Z is 1,2,3-triazol-5-yl, 2-methyl-1,3,4-thiadiazol-
5-yl, 2-methyl-1,3,4,oxadiazol-5-yl, l-N-methyltetrazol-
5-yl, 1-carboxymethyltetrazol-5-yl or l-carboxyethyl-
tetrazol-5-yl.
., .
- 16 -
~ . . .
~ 8~3S
The most preferred ~ompounds of this group
of the Formula II are those in which R is
Ar'-CH-CO-
Y
wherein Ar' is phenyl, p-hydroxyphenyl, 4-hydroxy-3,5-
dichlorophenyl, 3-chloro-4-hydroxyphenyl, o-, m- or
p-aminomethylphenyl, 2-thienyl, 3-thienyl, l-cyclohexenyl
or 1,4-cyclohexadienyl and Y is amino, hydroxy or
carboxy.
Another preferred subclass within this group
consists of the D-isomers of the acids in which R is
a-amino-a-(p-hydroxyphenyl)acetyl, a-amino-a-(3-chloroo
4-hydroxyphenyl)acetyl, a-amino-a-(3,5-dichloro-4-
hydroxyphenyl)acetyl ~-amino-a-(2-thienyl)acetyl,
a-amino-a-(3-thienyl)acetyl, a-amino-a-(l-cyclohexenyl)-
acetyl, a-amino-a-(1,4-cyclohexadienyl)acetyl, a-hydroxy-
acetyl, a-amino-a-(2-thienyl)acetyl, a-hydroxy-a-
(3 thienyl)acetyl, a-hydroxy-a-(l-cyclohexenyl)acetyl
- 17 -
or ~-hydroxy-~-(1,4-cyclohexadienyl)acetyl, or pharma-
ceutically accepta~le salts thereof.
The present invention further provides var-
ious novel intermediates useful in the synthesis of
the 7-acylamido 0-2-isocephem compounds of formula II
described above. .
Preferred embodiments of the present inven-
tion are the novel intermediates having the formula
H H
O
:~ ~ N ~
O I CH2-S-Z
C2R"
IV
wherein Z is optionally substituted Cl-C6 alkyl, aryl,
aralkyl or heterocyclic and R" is hydrogen or an
. easily cleavable ester carboxyl-protecting group, and
salts thereof. The preferred Z substituents are as
defined above in connection with the compounds of
formulae I and II.
Other preferred embodiments of the present
invention are the intermediates having the formula
H2N~ _ --
f
~ ~ N ~
CO R~ CH2S Z
III
- 18 -
~ .
13S
wherein Z is optionally substituted Cl~C6 alkyl, aryl,
aralkyl or heterocyclic and R" is hydrogen or an easily
cleavable ester carboxyl-protecting group, and salts
thereof. The preferred Z substituents are those mentione~
above as beinq preferred in connection with the compounds
of formulae I and II.
The intermediates of formulaeIII and IV may
be in the form of the free carboxylic acid or a salt
thereof or in the form where the carboxyl group is pro-
tected in a conventional manner such as preferably by
esterification. ~he protecting group is selected so that
it may be removed by methods which do not result in any
appreciable destruction of the remaining portion of the
molecule. Preferred carboxyl protecting groups are the
easily cleavable esters as defined above including in
particular benzhydryl, p-nitrobenzyl, trichloroethyl,
silyl including especially trimethylsilyl, phenacyl, p-
methoxybenzyl, acetonyl, (lower)alkyl such as methyl, t-
butyl or ethyl, benzyl, triphenylmethyl, methoxymethyl,
acetoxymethyl, phthalidyl, indanyl and pivaloyloxymethyl.
The novel 7-acylamido compounds of formula II
may be prepared by N-acylating a 7-amino intermediate of
the formula H H
H N
2 ~ \
~ N ~
O ¦ CH -S-Z
C2R" 2
III
- 19 -
~3 ~
wherein Z is optionally substituted ~l-C6 alkyl, aryl,
aralkyl or heterocyclic and R" is hydrogen or an easily -~
cleavable ester carboxyl-protecting group, or a salt
thereof, with an acylating acid of the formula
R-COOH
wherein R is an organic acyl group, or with its functional
equivalent as an acylating agent for a primary amine
and, if desired, (a) when R" is a carboxyl-protecting
group, converting the 7-acylated ester to the free
acid compound or a physiologically hydrolyzed ester
or a pharmaceutically acceptable salt of said acid
or ester, or (b) when R" is hydrogen, converting the
7-acylated carboxylic acid to a physiologically
hydrolyzed ester or a pharmaceutically acceptable
salt of said acid or ester and, if desired, resolving
a resulting isomer mixture into its component isomers.
The 7-amino starting materials of general
: formula III are of use primarily as intermediates in
preparing the pharmacologically active N-acyl deriva-
tives of formula II. The free acids, physiologically
hydrolyzed esters and pharmaceutically acceptable salts
of said acids and esters of formula III, however, do
possess some antibacterial activity per se against
various pathogenic microorganisms.
~, .
.
'
~ ~ .
- 20 -
~ .
li~B135
The 7-acylamido 0~2-isocephem compounds cf
formula II are prepared by N-acylation according to known
methods of the 7-amino group of intermediate III with an
acylating acid of the formula
R-COOH
wherein R is an acyl group, or with its functional equi-
valent as an acylating agent for a primary amino group.
The acylating agents for preparing the products of formula
II are known, readily preparable by known methods or
described herein as well as in our Belgian Patent 837,265.
Intermediate III may be acylated either in the
form of the free carboxylic acid (or salt thereof) or as
an easily cleavable ester (or acid addition salt thereof).
Preferred esters include benzhydryl, benzyl, p-nitrobenzyl,
trichloroethyl, silyl (especially trimethylsilyl), phenacyl,
p-methoxybenzyl, acetonyl, (lower)alkyl including partic-
ularly methyl, ethyl and t-butyl, triphenylmethyl, methoxy-
methyl, acetoxymethyl, pivaloyloxymethyl, phthalidyl and
indanyl. The procedures for preparing esters of carboxylic
acids are disclosed in the literature and are well-known
to those skilled in the art of penicillin and cephal-
~ osporin chemistry. Methods for preparing certain of the
; more preferred easily cleavable esters, i.e. the pival-
oyloxymethyl, acetoxymethyl, methoxymethyl, acetonyl and
~.
- 21 -
. '
13~
phenacyl esters, are disclosed in U.S. Patent 3,284,451
and in U.K. Patent 1,229,453. Preparation of phthalidyl
esters of penicillins and cephalosporins is described
in South African ~atent Applications 72/3799 and 72/3800
The free acid form of intermediate III may also be
converted to a silyl ester, e.g. trimethylsilyl ester,
as by the methods described in the literature, e.g.
U.S. Patent 3,249,622. The silyl ester carboxyl-
protecting group may be easily removed following the
acylation reaction by hydrolysis or alcoholysis.
Prior to the acylation reaction, any re-
active substituents on the acylating acid or derivative
thereof, e.g. hydroxy, carboxyl or mercapto, may be pro-
tected by use of suitable protecting or blocking groups
which are well-known to those skilled in the art of ~-
lactam chemistry, e.g. as by acylation or silylation.
When the acylating agent contains an amino functional group
in the acyl moiety, the amino group is protected by a
conventional amino-blocking group which may be readily
removed at the conclusion of the reaction. Examples
of suitable amino-protecting or blocking groups include
t-butoxycarbonyl, carbobenzyloxy, 2-hydroxy-1-naphth-
carbonyl, trichloroethoxycarbonyl, 2-ethoxycarbonyl-
l-methylvinyl and 2-methoxycarbonyl-1-methylvinyl. A
particularly valuable amino-blocking group is a proton,
- 22 -
~J~35
as in the acylating agent of the formula
~3 fH -C-c 1
NH2 HCl
Preferred amino-protecting groups are t-butoxycarbonyl,
carbobenzyloxy, the proton and a ~-diketon or
B-ketoester as in l~.K. Patent 1,123,333 or U.S. Patents
3,325,479 and 3,316,247, e.g. methyl acetoacetate, or
a ~-ketoamide as in Japan 71/24714. When the t-butoxy-
carbonyl, carbobenzyloxy, ~-ketoester, ~-diketon or
; ~-ketoamide protecting groups are employed, it is
preferred to convert the acylating acid containing
the blocked amino group to a mixed anhydride, e.g.
with ethyl or isobutyl chloroformate, before reaction ~ :
with compound III or a salt thereof. After the acylation
coupling reaction, the amino-protecting group and any
other functional protecting groups used may be removed
by methods known ~ se to form the desired product of
formula II. With respect to amino-protecting groups,
the t-butoxycarbonyl group may be removed by use of
formic acid, the carbobenzyloxy group by catalytic
hydrogenation, the 2-hydroxy-1-naphthcarbonyl group by
acid hydrolysis, the trichloroethoxycarbonyl group by
.~
:' :
.~
-- 23 --
.~ 8~5
treatment with zinc dust in glacial acetic acid, ~he
proton by neutralization, etc.
Acylation of a free amino group of a cephalo-
sporin or penicillin nucleus is a well-known reaction, and
any of the functional equivalents of the carboxylic acid
RCOOH ~ommonly used in penicillin or cephalosporin
chemistry as acylating agents for primary amino groups
may be employed in acylating intermediate III. Examples
of suitable acylating derivatives of the free acid include
the corresponding acid anhydrides, mixed anhydrides
(e.g. alkoxyformic anhydr~des), acid halides, acid azides,
active esters and active thioesters. The free acid
may be coupled with compound III after first reacting
said free acid with N,N'-dimethylchloroformininium
chloride [cf. Great Britain 1,008,170 and Novak and
Weichet, Experientia XXI, 6, 360(1965)] or by the use
of enzymes or of an N,N'-carbonyldiimidazole or an
N,N'-carbonylditriazole [cf. South African Specification
63/26843 or a carbodiimide reagent [especially N,N'-
dicyclohexylcarbodiimide, N,N'-diisopropylcarbodiimide
or N-cyclohexyl-N'-(2-morpholinoethyl)carbodiimide:
cf. Sheehan and Hess, J.A.C.S., 77, 1967 (1955)~, or
of alkylylamine reagent [cf. R. Buijle and H. G. Viehe,
Angew. Chem. International Edition, 3, 582, (1964)] or
- 24 -
~813~
of an isoxasolium salt reagent ~cf. R. B. Woodward,
R. A. Olofson and H. MAyer, J. Amer. ~hem. Soc., 83,
; 1010 11961)], or of a ketenimine reagent Icf. C. L.
Stevens and M. E. Munk, J. Amer. Chem. Soc., 80, 4065
(1958)l or of hexachlorocyclotriphosphatriazine or
hexabromocyclotriphosphatriazine (U.S. 3,651,050) or of
diphenylphosphoryl azide lDPPA; J. Amer. Chem. Soc., 94,
6203-6205 (1972)] or of diethylphosphoryl cyanide [DEPC;
Tetrahedron Letters No. 18, pp. 159S-1598)] or of
diphenyl phosphite ITetrahedron Letters No. 49, pp. 5047-
5050 (1972)]. Other examples of suitable amide coupling
reagents which have been described in the literature
include (CH3)2SCH2CCHBr/DMSO (J. Chem. Soc. (C) 1904
(1969), HCCOCH3 (Rec. Trav. Chim. 74 769 ~1955),
(CH3)2C(OCH3)2 (Chim. Ther. 2, 195 (1967), SiC14
(J. Org. Chem. 34, 2766 (1969), TiC14 (Can. J. Chem.
48, 983 (1970), (PNC12)3 (J. Org. Chem. 33, 2979 (1968)
SO3-DMF (J. ~. Chem. 24, 368 (1959), ion exchange
resins (Helv. 44, 1546 (1961) and J.C.S. C, 874 (1969)
and ~ Ie (J, Chem. Soa. 4650 (1964).
N
:,
- 25 -
8135
An equivalent of the acid chloride is the corresponding
azolide, i.e., an amide of the corresponding acid whose
amide nitrogen is a member of a guasi-aromatic five
mem~ered ring containing at least two nitrogen atoms,
i.e., imidazole, pyrazole, the triazoles, benzimidazole,
benzotriazole and their substituted derivatives. As an
example of the general method for the preparation of an
azolide, N,N'-carbonyldiimidazole is reacted with a
carboxylic acid in equimolar proportions at room temper-
ature in tetrahydrofuran, chloroform, dimethylformamide
or a similar inert solvent to form the carboxylic acid
imidazolide in practically quantitative yield with
liberation of carbon dioxide and one mole of imidazole~
Dicarboxylic acids yield dimidazolide. The by-product,
imidazole, precipitates and may be separated and the
imidazolide isolated but this is not essential. A
preferred acylating agent for preparing 7-acylamido
~; compounds containing an ~-amino substituent, e.g.
~-aminobenzyl, ~-amino-~-thienylmethyl, etc. is the
- 20 N-carboxy anhydride (Leuch's anhydride). In this
structure the group which activates the carboxyl
group also serves to protect the amino group. Another
` preferred acylating agent for introducing a side
.
il~813~
chain containing an ~-amino functional group is the
acid chloride hydrochloride, of the formula
Ar-CH-COCl
NH2-HCl
which also serves a dual function of carboxyl activation
and amino protection. Mention was made above of the
use of enzymes to couple the free acid with compound
III. Included in the scope of such processes are
the use of an ester, e.g. the methyl ester, of that
free acid with enzymes provided by various micro-
organisms, e.g. those described by T. Takahashi et al.,
J.A.C S , 94(11), 4035-4037 (1972) and by T. Nara et
al., J. Antibiotics (Japan) 24(5), 321-323 (1971) and
in U.S. 3,682,777. A particularly preferred coupling
agent for coupling the acylating acid with compound III
(or a salt or ester thereof) is N-ethoxycarbonyl-2-
ethoxy-1,2-dihydroquinoline (EEDQ) as described in
J A.C.S., 90, 823-824 and 1652-1653 (1968) and U.S.
Patent 3,455,929.
The particular process conditions, e.g. tem-
perature, solvent, reaction time, etc. selected for
the coupling reaction are determined by the nature of
the reactants and acylation method used and are known
to those skilled in the art.
- 27 -
;, ' '''''' ~-- ~'
The acylating agents which may be used to form
the N-acyl compounds of formula II are known in the
literature along with methods fc,r their synthesis or
are disclosed in the examples which follow. In those
cases where the acylating agent contains one or more
asymmetric carbon atoms and thus exists in optically
active forms, the compounds obtained using such an
acylating agent are ordinarily obtained in racemic
form. When the separate optical isomers are desired,
the acylating agent can be resolved in a conventional
manner such as by reacting the free acid with cinchonine
strychnine, brucine or the like, fractionally crystal-
lizing to separate the diastereoisomeric salts and
separately acidifying the solid phase and the liquid
phase to liberate the optical isomers.
The 7-acylamido compounds of the present in-
vention may be isolated in any of the ways customarily
employed for the isolation of corresponding cephalosporin
compounds. Formation of a desired pharmaceutically
acceptable carboxylic acid or acid addition salt is
carried out by known methods, e.g. reaction of the
acid of compound II (or ester in the case of acid
addition salts) with an appropriate base or acid.
- 28 -
13~
A compound of ormula lI in the form of the free
acid or a salt thereof may be converted to a pharma-
ceutically acceptable ~al~ thereof or to a physiologic-
ally hydrolyzed ester or pharmaceutically acceptable
salt thereof. Similarly, the product of formula II
in the form of an easily cleavable ester or salt thereof
may be converted to the free acid product or a pharma-
ceutically acceptable salt thereof by removal of the
esterifying group to form the free acid, e.g. by acidic
or alkaline hydrolysis, by enzymatic hydrolysis (as
with human or animal serum), by hydrogenolysis or by
treatment with chemical reagents known to remove partic-
ular blocking groups, e.g. sodium thiophenoxide as in
U.S. 3,284,451, and subsequent treatment of the free
acid with an acid or base to form a pharmaceutically
acceptable salt.
The easily cleavable esters of the compounds
of formula II are useful as intermediates in the produc-
tion of the free acid product. The pivaloyloxymethyl,
acetoxymethyl, phthalidyl, indanyl and methoxymethyl
esters are also useful as active antibacterial agents
since on oral administration they are rapidly hydrolyzed
to the active metabolite. These esters are of partic-
ular interest because they provide on oral administration
different rates and amounts of absorption and give dif-
fering concentrations of the active antibacterial agent
in blood and tissues.
- 29 -
3135
The 7-amino intermediates of general foxmula
III may be prepared by selectively reducing a 7-azido
intermediate of the formula
H H
N3
O CH~S-Z
C2R'
IV'
wherein Z is optionally substituted Cl-C6 alkyl, aryl,
aralkyl or heterocyclic and R" is an easily cleavable
ester carboxyl-protecting group. The carboxyl-
protected compound may, if desixed, be cleaved to
produce the free-acid intexmediate III which can be
converted to a salt by methods known per se.
Preferred reducing agents for use in pre-
paring the intermediates of formula III include chemical
reducing agents such as zinc and ammonium chloride,
aluminum amalgam and hydrogen sulfide in the presence
of a base, e.g. triethylamine or ammonia. Catalytic
hydrogenation may also be employed with such catalysts
as noble metals, preferably platinum or palladium
including derivatives thereof such as oxides, hydroxides,
and halides, or Raney nickel, said catalysts being option-
ally supported on a conventional carrier such as carbon or
diatomaceous earth. Catalytic hydxogenation is performed
~ .
- 30 -
~ ~)~35
with a non-reducible inert solvent, e.g. methanol,
ethanol or ethyl acetate, and preferably at
atmospheric or slightly ele~ated pressure at room
temperature.
Compound III in the carboxyl-protected form
or a salt thereof may be used directly as a starting
material in the N-acylation process discussed above.
Alternatively, the protected intermediate may be de-blocked
to form the free carboxylic acid which may then be option-
ally converted to a salt or to another carboxyl-protected
form, e.g. a physiologically hydrolyzed ester or salt
thereof. By proper selection of reduction conditions
and protecting groups, azido intermediate IV' may be
converted either simultaneously or in stepwise fashion
to the 7-amino free acid III. Thus, if mild hydro-
genation conditions are used, e.g. catalytic hydro-
genation with 10% Pd-on-charcoal or a mild chemical
reducing agent such as H2S in the presence of a base such
as triethylamine or ammonia, the azido group may be
reduced without concomitant removal of esters resistant
to such conditions, e.g. benzyl or p-nitrobenzyl. If
stronger reducing conditions are used such as 30%
Pd-on-diatomaceous 2arth, both the azido group and
most reducible esters will be simultaneously reduced.
.~
' :
- 31 -
~ 8~3~
A preferred embodiment of the present invention
is the process comprising the consecutive steps of
1) selectively reducing a 7-azido intermediate
of the formula IV' to produce a carboxyl-protected 7-
amino intermediate of formula III and/ if desired,
removing the carboxyl-protecting group to produce
the corresponding free acid intermediate of formula
III or optionally a salt thereof; and
2) N-acylating intermediate III or a salt
thereof with an acylating acid of the formula R-COOH
where R is an acyl group/ or with its functional
equivalent as an acylating agent for a primary
amine and, if desired, (a) when R" is a carboxyl-
protecting group, converting the 7-acylated ester
to the free acid compound or a physiologically
hydrolyzed ester or a pharmaceutically acceptable
salt of said acid or ester, or (b) when R" is
hydrogen, converting the 7-acylated carboxylic acid
to a physiologically hydrolyzed ester or a pharmaceuti-
cally acceptable salt of said acid or ester and, if
desired, resolving a resulting isomer mixture into
its component isomers.
- 32 -
~3~
The 7~-azido intermediates IY' may be pre-
pared by two alternative methods. In one procedure a
dihalide intennediate of the formula
H H
C ~X) (CH2X' )
C2R ~
V
wherein Y' represents a displaceable leaving group,
preferably a group such as halo or sulfonyloxy,
; e.g., alkyl- or substituted alkylsulfonyloxy or
aryl- or substituted arylsulfonyloxy, and most
preferably a group selected from halo, -OSO2-
(lower)-alkyl including especially -OSO2CH3, -OSO2CF3
and -OSO2C6H4CH3(para), X and X' which may be the
same or different each represent a halogen atom,
preferably bromine or iodine and most preferably
iodine and R' is an easily cleavable ester carboxyl-
protecting group is reacted in an inert organic
solvent in the presence of an acid acceptor with a
nucleophile of the formula
Z-SH
wherein Z is optionally substituted Cl-C6 alkyl,
aryl, aralkyl or heterocyclic, or a salt thereof,
:'
- 33 -
~ .
to form a thiolated intermediate of the formula
Y' ., .
H H
N3 - J
~N~C (X) (CH2-S-Z)
C02R
VI
Intermediate VI is then converted to the desired 7B-
azido compound by cyclization with base in an inert
: organic solvent.
The dihalide starting material V may be used
in either of its isomeric forms
~ , .
H H ¦
N3 _ _ )
~ X
CH2X'
2R '
Va
or
Y'
H H
: N ~ CH2X'
.' I I
.~ ~ N
O
C2R'
, Vb '
- 34 -
s
or as a mixture of isomers. Formula V above is in-
tended to represent either of the individual isomers
or the mixture. Any dihalide including a mixed
dihalide, e.g. X ~ Cl, X' - Br, may be used but the
most preferred compound is the diiodide. Compound V
is reacted in an inert organic sol~ent, e.g.,
methylene chloride, with the desired thiol or a salt
thereof. Since an acid HX is given off during the
reaction, an acid acceptor, preferably an organic
base such as pyridine or a trialkylamine, is used.
Compound V, the thiol and the acid acceptor are
preferably employed in approximately equimolar
amounts. The temperature for the displacement step
is not critical, but best results are obtained at
room temperature or below, most preferably at a
temperature of about 0C.
Cyclization of intermediate VI is carried
out in an inert organic solvent, preferably a polar
organic colvent such as dimethylsulfoxide or dimethyl-
formamide, with a suitable base. The base used in
the cyclization step may be selected from a wide variety
of bases including especially those of the following
categories:
(a) anion~ derived from carboxylic acids having
a PKa Of between about 3.5 and 5.5;
- 35 -
.. - ' - :
3L~.'D8135
~b) tertiary organic amines such as a trialkyl-
amine (e.g., triethylamine~, pyridine,
n-methylpiperidine, n-methylmorpholine, etc.;
(c) alkali metal hydrides, e.g., sodium or
potassium hydride; and
(d) organolithium compounds including especially
lithium alkyls, e.g., met~yl, lithium or
butyl lithium.
Most preferred cyclization bases are acetate and
formate anions, e.g., from alkali metal, ammonium or
substituted ammonium formates or acetates. The most
preferred base is acetate anion. The base is pre-
ferably used in a molar excess relative to compound
VI. While the preferred temperatures for this step
are room temperature or below, the temperature is not
critical. The leaving group Y' in formula Vl should
be one which is efficiently displaced under the
conditions of the base cyclization reaction and is
preferably halo (chloro, bromo, fluoro, Dr iodo) or
sulfonyloxy. The most preferred leaving group is the
mesylate group.
Preparation of intermediate V is described
in the text below under the heading "Preparation of
Starting Materials". Briefly summarized, the reaction
scheme is as shown in Flow Sheet I below:
'
- 36 -
;- ~L~ 3~ ~
HON ~bo~yl ~oee tion ~s
y eta ~zat~on
C-C~CH3 ~ >
C02R '
HONO'~\ nitroso
~ reduction >
CO2R' CH3
H2N O ~ Schiff base
:~ ~ C ~ ~ form~tion as _ >
~ C02R' CH3 clnnamaldehyde
.; '
., .
lactam formation
as with
: ~o ~ azidoacetyl halide >
H3
C02R '
~ . ,
H H
`~ N ~ -
ozonolysis
C~ ~c ' ' '
CO2~' 3
- 37 -
:' 1
'
~ 31 3
N3 _ 6elective aldehyde
~/ 0~ , reduction >
.. ~ N
C02R ~
H H CH OH conver~ion of
N3 - ~ 2 hydroxyl group to a
~ore activ~ted _
I u ~ leaving group as by
~ , N ~ O halogenation or
~ ~H3 esterification with
CO2R' a ~ulfonic scid derivative
H H ISO2CH3
3 ~ de-ketalization >
~N ,~ o
CO2
OSO CH hydroxyl activation
N H H 1 2 3 ns with triflic
3 ~ snhydride >
N
C02R '
N H ~ 2 3 ~llene form3tion
3 - - J ~ith bsse >
OS02CF3
H3
C2R '
-38-
~8
H H J 2 3
halo~enation , ~ V
~,~C--CH2
~02R l
Flow Sheet 1
- 3 9 -
81:~S
An alternative method ~or preparing the 7~-
a~ido intermediates of formula IV' involves reacting
an intermediate of the formula
H H
N
r~N I
' 0~ \~CH2Y"
2
VII
wherein Y" represents a displaceable leaving group
and R~ is an easily cleavable ester carboxyl-pro-
tecting group in an inert organic solvent in the
; presence of an acid acceptor with a thio of the
formula
HS-Z
wherein Z represents an optionally substituted
Cl-C6 alkyl, aryl, aralkyl or heterocyclic group,
or a salt thereof.
Leaving group Y" in formula VII may be
any nucleophilic group which is displaceable by the
thiol. Examples of suitable leaving groups include
halo (chloro, bromo, fluoro or iodo) and sulfonyloxy,
i.e., alkyl- or substituted alkylsulfonyloxy or aryl-
or substituted arylsulfonyloxy. Preferred leaving
groups are halo, -OSO2-(lower~alkyl including
especially -OSO2CH3, -OSO2CF3 and -OSO2C6H4CH3(para).
A most preferred leaving group is the mesylate group.
- 40 -
~1~8135
The nucleophilic displacement reaction is
conducted in an inert organic solvent, e.g., methylene
chloride r in the presence of an acid acceptor, pre-
ferably an organic base such as pyridine or a trialkyl-
amine. ~he heterocyclic thiol, acid acceptor and
intermediate VII are preferably used in approximately
equimolar quantities. The temperature is not critical,
but best results are obtained at room temperature or
below, most preferably at a temperature of about 0C.
The preferred starting materials of formula
VII used in the above process include Y" = halo or sulfonyl-
oxy .
A preferred method of preparation of these
starting materials involves the steps of
(1) cyclizing in an inert organic solvent,
e.g., dimethylformamide, a dihalide inter-
mediate of formula V with a base R2COO
in which R2 in hydrogen or (lower) alkyl
to produce an intermediate of the formula
H H
N3 _ _ " ~~
~ l '
~ N ~ H2OCOR2
C 2R'
:',
in which R2 is hydrogen or (lower) alkyl and;
.
,,.,~
- 41 -
813S
~2) subjecting said intermediate to acid
hydrolysis to produce an intermediate of
the formula
H H
N3 _
~ l
; o ~ CH2 ; and
C2R '
either
(a) esterifying the 3-hydroxymethyl intermediate
with a sulfonic acid derivative, e.g.,
methanesulfonyl chloride, p-toluenesulfonyl
chloride or triflic anhydride, in the pre-
sence of an organic base and in an inert
organic solvent, e.g., methylene chloride,
to produce the desired 3-sulfonyloxy deriva-
tive of formula VII; or
(b) halogenating the 3-hydroxymethyl intermediate
with a phosphorus halide, e.g., phosphorus
trichloride, or phosphorus tribromide, to
produce the desired 3-halomethyl derivative
of formula VII.
- 42 -
~L3~ii
The present invention also provides an alter-
native method for preparing the novel 7-acylamido
compounds of formula II; which process comprises
reacting in an inert organic solvent in the pre-
sence of an acid acceptor a 7-acylated compound of
the formula
H R
R-NH ~
~
~N~` CH2Y"
C02R"
VIII
wherein R is an acyl group, Y" is a displaceable
leaving group and R" is hydrogen or an easily
cleavable ester carboxyl-protecting group, or a
salt thereof, with a thiol of the formula
HS-Z ~:
wherein Z is optionally ~ubstituted Cl-C6 alkyl,
aryl, aralkyl or heterocyclic, or a salt thereof
and, if desired, (g) when R" is a carboxyl-protecting
group, converting the 7-acylated ester to the free
acid compound or a physiologically hydroylzed ester
. ;,
~` or a pharmaceutically acceptable salt of said acid
or ester, or (b) when R" is hydrogen, converting the
- 7-acylated carboxylic acid to a physiologically
hydrolyzed ester or a pharmaceutically acceptable salt
of said acid or ester; and, if desired, resolving a
resulting isomer mixture into its component isomers.
- 43 -
The nucleophilic displacement of the Y" group
in compound VIII may be carried out using the same
general reaction conditions, i.e., solvents,
temperature range, acid acceptors, as described
above in connection with thiolation of compound V.
The 7-acylated starting materials for
the above process may be prepared by the procedures
disclosed in the art.
While Y" may be any nucleophilic leaving
group which is displaceable by the desired thiol,
the preferred starting materials are compounds
wherein Y" is halo (chloro, bromo, iodo or fluoro)
or sulfonyloxy, i.e., alkyl- or substituted alkyl-
sulfonyloxy or aryl- or substituted arylsulfonyloxy.
Examples of preferred leaving groups include halo,
-S2 -(lower)alkyl, -OSO2CF3 and -OSO2C6H4CH3(para).
- A most preferred leaving group is the mesylate
group.
Starting materials of formula VIII may be
prepared from 73-azido compounds of formula VII by
the consecutive steps of
(1) Selectively reducing intermediate VII
as by catalytic hydrogenation with a
noble metal catalyst, e.g., 10~ Pd-on-
charcoal, or by the use of chemical
reducing agents such as zinc and ammonium
chloride, aluminum amalgam or hydrogen
~ '
- 4~ -
~813~
sulfide in the presence of a base ~uch
as ammonia or triethyl2mine to pro-
duce a 7-amino intermediate of the
formula
H H
: 2
f
/ N ~ CH2-Y" ; and
CO R'
(2) N-acylating said 7-amino intermediate or
a salt thereof with an acylating acid of
the formula
R-COOH
!,
~ wherein R is an acyl group, or with its functional
: equivalent as an acylating agent for a primary amine,
, and, if desired, converting the carboxyl-protected
7-acylamino compound to the corresponding free acid
or to a salt thereof as by the methods disclosed
above in connection with the de-blocking of compound
II.
`,''
,
- 45 -
13~
The pharmaceutically active compounds of
the present invention are potent antibacterial agents
useful in the treatment of infectious diseases in
poultry and animals, including man, caused by many
Gram-positive and Gram-negative bacteria. The active
compounds are also of value as nutritional supplements
in animal feeds and as agents for the treatment of
mastitis in cattle.
The novel medicaments provided by the pre-
sent invention may be formulated as pharmaceutical
compositions comprising, in addition to the active
ingredient, a pharmaceutically acceptable carrier or
diluent. The compounds may be administered both orally
and parenterally. The pharmaceutical preparations may be
in solid form such as capsules, tablets or dragees, or in
liquid form such as solutions, suspensions or emulsions.
In the treatment of bacterial infections in man, the active
compounds of this invention may be administered parenterally
or orally in an amount of from about 5 to 200 mg~Kg./day
and preferably about 5 to 20 mg./Kg./day in divided dos-
age, e.g. three or four times a day. They are administered
in dosage units containing, for example, 125, 250 or
500 mg. of active ingredient with suitable physiologi-
cally acceptable carriers or excipients.
.
- 46 -
~13~ii
Illustrative examples of the preparation of
starting materialS and compounds of the present
invention follow. These examples are given in
illustration of, but not in limitation of, the
present invention. All temperatures are in degrees
Centrigrade. DMF represents dimethylformamide, THE
stands for tetrahydrofuran and EEDQ is the amide
- bond forminq reagent having the structure
OEt
;~ CO2Et
'.:
The ~-lactam compounds prepared in the
-., 10 examples which follow all have the hydrogen atoms at
,. carbons 6 and 7 cls with respect to each other and,
unless indicated, the products are racemic mixtures
; in the sense that they are composed of equal parts
of the two isomers having the following structures:
H H H H
>N~ ,~N<
", C--O-- C--O--
::
.
- 47 -
1~813S
pREPARATION OF STARTING MATERIALS
. _ . . .
Preparation 1
A. 2 3 OS 2 3
H H H H
3 ~ Br ~ -C~32Dr
N ~ CD2Dr ~ Dr
C02CH20 02CH20
Benzyl Oximino-Acetoacetate
CH3C - Cl - CO2CH20
O NOH
1.1
The procedure was essentially the same as that
used to make the corresponding ethyl ester by H. Adkins
and J. Reeve, JACS 60, 1328 (1938).
In a three necked one liter flask fitted with
a thermometer, a dropping funnel and a magnetic stirrer
; were placed 173 g. (0.9 mole) of benzyl acetoacetate
lThe benzyl acetoacetate was prepared as described by
Baker et al., J. Org. Chem. 17, 91 (1952)] and 130 ml.
of glacial acetic acid. The contents were cooled in an
ice bath and a solution of 69 g. (1 mole) of sodium
- 48 -
11~J1813~
nitrite in 130 ml. of water ~as added over a period
of half an hour; the temperature was kept at 0 to
10C. After the reaction mixture was stirred for
one hour at room temperature, 400 ml. of water was
added and the stirring was continued for an additional
two hours. The reaction mixture was extracted three
times with 200 ml. portions of diethyl ether. The
diethyl ether extracts were combined, washed once
with water, three times with saturated sodium bicar-
bonate solution and once with brine. After drying
over anhydrous sodium sulfate, the diethyl ether solu-
tion was evaporated leaving [1.1] as a clear oil which
solidified upon trituration with petroleum ether
(30-60) to give 186.5 g. (93.2%) of white solid.
Its NMR spectrum was consistent with the assigned
structure. Generally the product was used as such
in subsequent reaction but it can be recrystallized
from toluene, m.p. 81-82C.
_nzyl Oximino-Acetoacetate Ethylene Xetal
CH3CI~ CO2CH20 ketalization~ CH3 -~C~ Cl CO2CH20
o NOH ~ NOH
1.1 2.1
:'
In a two liter flask fitted with a Dean
Stark water separator and a condenser were placed 186.5 g.
.
- 49 -
8~3~
(0.85 mole) of benzyl oximino-acetoacetate ~ , 62 g.
(1 mole) of ethylene glycol, 800 ml. of benzene (reagent
grade) and 2 g. (10.5 m~ole) of p-toluenesulfonic acid
monohydrate. The reaction mixture was boiled at reflux
~ntil 15 ml. of water was removed (3 houIs). ~he benzene
solution was washed once with saturated sodium bicar-
bonate solution and once with brine. After drying over
anhydrous sodium sulfate, the benzene solution was
evaporated, leaving 212 g. (94% yield) of benzyl oximino-
acetoacetate ethylene ketal 12.1] as a light yellow oil!
Its NMR spectrum was consistent with the assigned
structure. Generally, the product was used as such
in subsequent reactions but one of the isomers can be
crystallized2 from toluene-petxoleum ether (b.p. 30-
60C.); m.p. 52C.
Anal- Calc'd- for C13H15N5 C, 58-86; H~ 5.70; N~ 5-28-
Found: C, 58.97 ~, 5.68: N, 5.12.
1. A mixture of the syn and anti isomers.
2. Only 35% of the oil could be crystallized.
-- 50 --
1~08~3~
Benzyl Amino-Acetoacetate Ethylene Ketal
Selective ~ -
reduction ~
CH3/C\- 11 - CO2CH20 CH3/ ~~ FH ~ C2CH20
NOH ~__J 2
2.1 3.1
Freshly prepared aluminum amalgaml (from 27 g.
of aluminum foil) in a three-necked one liter flask was
, covered with 500 ml. of diethyl ether. The flask was
fitted with a mechanical stirrer, a condenser, and a
dropping funnel. A solution of benzyl oximino-aceto-
acetate ethylene ketal [2.1] (132.5 g.; 0.5 mole) in
300 ml. of wet diethylether2 was added dropwise at such
a rate as to maintain boiling at reflux. After stirring
for four hours, the reaction mixture was filtered
through a Buchner funnel. The filtrate was evapor-
ated leaving 110 g. of yellowish oil. The oil was
dissolved in 800 ml. of dry diethylether and dry
hydrogen chloride gas was bub~led into the solution
~` 20 until no further precipitation occurred. The white
; precipitate was filtered off and washed once with
diethylether and then dried in vacuo. This provided
j 108 g. of benzyl aminoacetoacetate ethylene ketal
hydrochloride [3.1]; m.p. 157-158~C.
- 51 -
`:
.
3~
Anal. Calc'd. for C13H17N04-HCl: C, 54.26; ~, 6.31;
N, 4.87.
Found: C, 53.9~; H, 6.19;
N, 4.60.
To obtain the free base, the hydrochloride
salt was suspended in 500 ml. of diethylether and con-
centrated ammonium hydroxide was added with shaking
until the solid went into solution. The diethylether
layer was separated and washed twice with brine. After
drying over anhydrous sodium sulfate, the solvent was
; evaporated leaving 90 g. (71% yield) of colorless oil.
1. The aluminum amalgam was prepared essentially as
described in A.I. Vogel ("Practical Organic
Chemistry", 3rd, Edn., Longemans Green & Co.,
London, (1957), p. 198) except for the following
modification:
(a) 5% NaOH was used.
(b) The second washing with ethanol was
omitted.
.
- 52 -
.,,, ., ..
~Q ~3
(c) Dry diethyle~her was used for washing snd
m2st of the water must be d~ained.
2. ~he d~ethylether was saturated with water by sh~king
with water in 8 separato~y funnel.
3. The product can be stored as the hydsochloride salt.
Schiff Base Formation And B-Lactam Form2tion
' 0
; ~ Schiff base ~O
~ ~ formation ~ N~ ~
CH ~ - CH - C0 CH20 ~ CH3
:` NH2 C02CH20
3.1 ~ ~ 4.1
H H
0 ~ 3
~2CH20
5.1
Sn ~ one litc flask fitted with a Dean Stark wate:
: separator and a condenser were placed 70.3 g. (0.28 mole)
,
: benzyl aminoacetoaceta~e ethylene ketal t3.1], 37 g. (0.2a
~ mole) cinnamaldehyde, a~d 750 ml. of methylene chlorid~
a3
.
~Q8~35
(reagent ~radel. The mixture ~as boiled at reflux for
30 minutes and then 400 ml. of methylene chloride
were distilled off. The concentrated solution was
then dried over anhydrous sodium sulfate and then eva-
porated to dryness in vacuol. The residual oil was
checked by NMR to ensure that Schiff base formation
was complete before continuing on to the next step.
The freshly prepared Schiff [4.1~ was diluted
with 600 ml. of methylene chloride2 and cooled to 02C.
(ice-salt bath). Triethylamine (31.1 g.; 0.308 mole)
was added and then a solution of 36.2 g. (0.308 mole)
of azidoacetyl chloride3 in 362 ml. of methylene
chloride2 was added dropwise at 0C. over a period of
one hour. The reaction mixture was stirred for an
additional hour at room temperature4 and then evapor-
ated on a rotary evaporator at reduced pressure while
being heated on a 35C. water bath5. The residue was
diluted with 500 ml. of diethylether and filtered.
The filtrate was washed twice with brine and dried
over anhydrous sodium sulfate. Evaporation of this
solution yielded 117.5 g. (94% yield) of styryl
~-lactam[5.1]. Its NMR and IR spectra are consistent
with the assigned structure and indicate the presence
of a mixture of isomers, diasteriomeric at the carbon
to the carbonyl of the benzyl ester.
1. This evaporation must be done to ensure complete
- 54 -
Schif~ ~ase ~orm~tion.
2. All the me~hylene chlorlde used $n the cyclo-
addition reaction was re~gent gr~de which was
first dried over molecular sieve (Type 4A) and
then over anhydrous calcium chloride. ~t ~ -
storet thereafter over molecular sieve (Type
4A) . .
3. J. H. Boyer and J. Horner, J. Amer~ Chem. Soc.,
(1955), 77, 951.
4. ~he reection mixture can be ~ept overnite at
0 if necessary.
5. This operation is necessary to ensure complete
~-lactam formation.
H H
Cd3 2.(CH3)25 ~ OCH3
C02C~2
; 5.1 6.1
Styryl ~-lactam 15.1] (117.5 g.; 0.262 mole~ was
dissolved ln one liter of methylene chloride (reagent
srade), cooled to -50 to -60C. in a dry ice-acetone
bath, and ozonized until a faint blue-green color
appeared. ~he solution was then flushed with nitrogen
until the color faded. Methylsulfide tl ml.) was
-55-
3 ~ i
~dded to the -50C. solution, which was then allowed
to 610wly reach 25 as the cooling bath gradually -
melted. ~t was kept overnite at room temperature
under nitrogen and then it was washed twice with
1% ~od~um bicarbonate solution, twice with brine,
driet over anhydrous sodium sulfate, and evaporated
to dryness. ~he resulting oil triturated four
t~mes with 100 ml. portions of petroleum ether (b.p.
30-60C.) o remove benzaldehyde. The oil was then
triturated carefully with diethylether whereupon
lt solidified. The solid was filtered off and
; dried to provide 75 g. (71.5%) of aldehyde 16.1] as a
m~xture of isomers diasteriomeric at the carbon a
to the carbonyl of the benzyl ester. Recrystalliza-
tlon of [6.1] from ether ~ave white crystals, m.p.
101-102C. (co~rected).
~nal. Calc'd. for C17H18N406 : C, 54.54; H, 4-84;
N, 14.96.
Found : C, 54.75; H, 4.87;
` N, 14.89.
-56-
N ~ O ~l ~ }~
3 `ff' /--~ Na~4 3 ~J,~
d~><~3 T~F o~ ~3
CO2CH20 o C02CH2~
7.1
2 20
8.1
The ~ldehyde 16.1~ (116.3 g.; 0.31 mole) was d~ssolved
~n 600 ml. of T~F (reagent gr~de) and the solution w8s
then cooled to -10C. (ice-methanol bath). Sodium
borohydride (5.88 g.; 0.155 mole) was added and the
reaction mixture was stirred 1 hour. 10% ~queous hydro-
chloric acid was added until the mixture was slight~y
~cidic, then 600 ~1. brine was added. The THF layer
was separated and the aqueous phase was extracted twice
w~th 250 ml. port~ons of diethyle~her. The combined
organic phases were washed twice wi~h 400 ~1. portions
of brine, dried over ~nhydrous sodlum sulfate, and
evaporsted ln va~uo to yield 117.3 g. of crude alcohol
-57-
[7.1l ~s an orange oil, This oil w~s used ~s 6uch in
the next rea~tion.
A solution of methanesulfonyl chloride ~37.8 g.;
0.34 mole) in 100 ml. of methylene chloridel w~s added
dropwise at 0C. (ice-water bath) to a stirring solu-
tion of alcohol ~7.1] (105.6 g,; 0.28 mole, triethyl-
a~ine (56.6 g.; 0.34 mole) and one liter of methylene
chloridel. A~terwards, the reaction was stirred for
30 hours at 25C, It was then washed twice wlth brine
(500 ~1. portions), dried oves anhydrous sodlum sulfate,
and eva~orated in vacuo The result~ng oil was dissolved
in methylene chloride, treated with norite, znd then
filtered over approximately 200 g. of activity I silica
gel. The silica gel was then washed with approximately
2 liters of methylene chloride. The filtrate was evap-
orated to dryness and the resulting oil (116 g.) was
covered with diethylether. It crystallized on stsnding
to give 87.2 g. (80% from ~6.1]~ of mesylate [8.1] as
off-white solid, m p, 97-99C. (corrected~
1. The methylene chloride used was reagent grade
which had been further purified by pass~ng over
8 column of calcium chloride and then storing
~; ~ver molecular sieve (Type 4A)
~a
~ 3~ '
- H H ~S02CH3
N3 ~ r~~ de-Ketal- ~ OH
I ¦ O O ization
N
~o2c~32es ~/ C02t:H20
8.1 ~ ~/ 9.1
, H H 2 3
3 ~ OSO2CF3
o ~ ~ H3
C02CH20
10.1
:,
A mixture o~ mesylate [8.1] (3.19 g.; 6.43 mmole)
and 30 ml. of 95% trifluoroacetic acid was stirred at
25 for 2 hours. The mixture was diluted with 300 ml.
of brine and extracted three times with methylene
chloride (100 ml. portions). The combined extracts
were washed three times with water (50 ml. portions,
unt~l neutral), dr~et (anhydrous so~iu~ sulfate) and
evaporated to dryness in vacuo lea~ing 3.17 g. of a
brown Dil. NMR spectra of this oil indicate the
presence of >90% enol (9.1].
Crude enol l9.1~ (48.0 g.; 0.~17 mole) and tr~fl~c
~nhydrlde (33.0 g.; 0.117 mole) were dissol~ed ~n 50n
~1. of methylene chlor~de ~nd the solution was the~
cooled to O-C. ~ce-water bath). A solution of trl-
ethylam1ne (11.8 g.; 0.117 ~ole) in 80 ml. of methylene
chloride2 was added d~opw~se o~er a per~od of 40 ~inutes.
When the additlon was complete, the ice-water bath was
removed and the mixture was stirred ~t 25 for 45 :: :
m~nutes. The ~ixture was then poured into 300 ~lo of
~ce WE~er and washed with colt water untll the pH of
the washings was approximately 6. The extract was
dr~ed (anhydrous sodlu~ sulfate) and ev~porated ~n vecuo ~ -
to give 54.0 g. of crude tr~late 1lO.l] as a dark sed :~
oil. This oil was dissolved in 400 ml. of benzene (US~)
~nd passed th~ough 2 1 1/2" pad of activity II~ s~lica
gel. The pad was washed with 1 1. of benzene. Evapor-
ation of the benzene gave 38.3 g. of a yellow oil. This
oil was carefully triturated with 50 ml. of absolute
ethanol and then cooled at 0C. for 2 hours. ~he
sesult~n~ whi_e solid was filte_ed off ~nd driet in
vac~ to give 19.5 g. o~ tr~flate ~1~.1] as one i~omer,
m~p. 57-59C. (corrected).
Anal. Calc'd. for C17l~l7F3N4O4S2: C, 37.67; H, 3.14;
~ , 10.33; S, 11.82.
Found: C" 37.40; H, 3.12;
N~ 10.43; S, 11.73.
~o
~ 3~'
1~ Triflic anhydride was prepared e~ follows:
170 g. (lOC ml.) CF3S03~ ( FLUOROCH~lIC ACID" - t- ademark -
3M Company) and 135 g. P2Qs were m~xed carefully,
shaken well, snd stored 18 hours protected fr~m
istu~e. The pro~uct was distilled from the
resulting sol~d m~8 us~ng a fla~e; the fraction
boiling 80-90~C. was collected. Re-dist~llation
of this fraction y~elded 119.45 g. (74%) of
tr~fl~c ~nhydr~de boiling 82-84C.
2. The methylene chloride used was reagent grade
lo which hsd been further purified by pass~ng over
a column of calcium chloride end then stored
over molecular s~eve (Type 4A).
H H. 2 3 H ~ S2~3
~: ~3 - - I orgsnic ~ N3 - J
~ OS02CF3 base ~ ~ ~ ~CH2
0 ~ ~ CH3 ~ ~ C
co2CH2~ ~ C02C~12P
H OS02CH3 H 0sa2CH3
+3 ~ ~ Br
C02CH2~ COZcH20~
~2.1 13.1
-61-
3~
Triethylamine (1 g.; 0.01 mole) was added to a
stirred solution of triflate llO.l] (5.42 g.; 0.01 mole)
in 55 ml. of methylene chloride (A.R.) at room temPera-
ture. After stirring for five minutes (at which point
TLC shows complete formation of allene [11.1~, a solution
of bromine (10 ml. of lM solution in CC14; 0.01 mole)
was added dropwise. After addition of the bromine,
the mixture was concentrated, absorbed onto Activity I
silica gel and dry column chromatographed on Activity I
silica gel by eluting with methylene chloride (VSP).
This yielded one fraction (unifoxmly one spot by TLC)
weighing 2.5 g. (45%). Its IR, W , and NMR spectra
were consistent with the expected dibromide structure
[13.1].
Anal. Calc'd. for C16Hl2Br2N4O6
N, 10.15.
Found: C, 35.25; H, 2.97;
N, 10.02.
- 6~ -
B . OSO~ CH 3
OSO CH
H H 1 2 3 H H
~ ¦ ~nd~ ~2I
O ~CH2I ~ I
C02CH20 C02CH20
A solution of triethylamine (101 mg., 1.00 mmole)
in 1.4 ml. of methylene chloride was added with stirring
to a solution of triflate from Preparation lA above
[10.1] (542 mg., 1.00 mmole) in 5.4 ml. of methylene
chloride at 0C. After allowing the solution to warm
to 24D over 15 minutes, a solution of iodine (254 mg.,
1.00 mmole) in 7.5 ml . of methylene chloride was added
with stirring over 30 minutes, then washed with water,
dried, decolorized, filtered and the solvent evaporated
in vacuo to give the diiodide [23.1] (588 mg.; 91%
yield) in greater than 95~ purity. The IR and NMR
spectra were consistent for the proposed structures.
Anal. Calc'd. for C16H16N406I2
N, 8.67; I, 39.28; S, 4.96.
Found: C, 29.76; H, 2.47;
N, 8.61; I, 39.37; S, 5.18.
- 63
Other suitable intermediates of general formula
V in the specification and claims may be prepared by:
(1) substituting another easilY cleavable ester
group for the benzyl ester of starting material
1.1; or
(2) esterifying compound 7.1 with another sulfonic
acid derivative or halogenating compound 7.1; or
(3) halogenatins allene 11.1 with a halogenating
agent other than bromine or iodine, e.g. BrCl. ~ -
Preparation 2
. ~
p-Nitrobenzyl 7~-azido-3-methylsulfonyloxymethyl-
~3-0-2-isocephem-4-carboxylate
H H
N3 ~ ~
N ~ CH20S02CH3
C2CH2 2
A solution of methanesulfonyl chloride (0.50 ml.,
6.5 mmole) in 10 ml . of methylene chloride was added
dropwise with stirring to a solution of p-nitrobenzyl
73-azido-3-hydroxymethyl- A3-o-2-isocephem-4-oarboxylate
;
- 64 -
~8~3~
(2.41 g., 6.43 mmole), triethylamine (0.97 ml., 7.0
mmole) and 75 ml. of methylene chloride at -10C. After
1/2 hour at -10 and 1 hour at 24, the ~olution was
washed with 5~ hydrochloric acid, 2% s~dium bicarbonate,
and water (85 ml. each), then the solvent was evaporated
in vacuo to give the mesylate title product, 2.86 g.
(98~ yield), as a yellow foam. The NMR of the product
was in agreement with the proposed ~tructure.
The p-nitrobenzyl 7~-azido-3-hydroxymethyl- ~3-
0-2-isocephem-4-carboxylate starting material used above
may be prepared as follows:
1. The diiodide intermediate of the formula
.'~ 0502CH3
~3 ~ Il
N ~ CH2I
Co2cH24~3No2
was prepared from p-nitrobenzyl acetoacetate according
to the procedures of Preparations lA and B (Starting
Materials) described above.
2. The diiodide intermediate (6.6 g., 9.6 mmole)
was cyclized with potassium formate (2.54 g., 30 mmole~
- 65 -
1~ 135
in a solution of 100 ml. DMF and 0.1 ml. water at 0.
After stirring for S hours with the cooling bath removed,
the mixture was poured into 100 ml. of cold water and
extracted with methylene chloride. After washing with
water containing a little NaCl, drying and evaporatiOn
in vacuo, p-nitrobenzyl 7~-azido-3-formyloxymethyl- ~ -
0-2-isocephem-4-carboxylate was recovered (5.3 g.) as
a brown oil.
3. To a solution of 5.3 g. of the 3-formyloxymethyl
intermediate in 53 ml. of acetone was added 26 ml. of
water and 3.2 ml. of 12M HCl. The mixture was stirred
at 24 for 7 hours, then poured into 100 ml. water and
extracted with methylene chloride. The combined extracts
were washed with water containing a little sodium
chloride, dried and evaporated in vacuo to give 3.6 g.
of a brown oil. The oil was absorbed from methylene
chloride onto 18 g. of silica gel and placed on a 72 g.
silica gel column (grade 3, 5% ether). The column
was eluted with 200 ml. of ether, then with ether/ethyl
acetate 3:1. The major component ~Rf 0.20) gave, on
evaporation of the solvent in vacuo, a yellow solid
which was recrystallized from acetone-ether to give the
3-hydroxymethyl starting material of this example, 950
mg. (17.5~ yield from the diiodide). m.p. 147-148.
''',
- 66 -
. .
11~ 135
Anal. Calc'd- for ClsH13N5O7: -
N, 18.66.
Found: C, 48.11; H, 3.61;
N, 18.81.
By replacement of the methanesulfonyl chloridein the procedure above with other sulfonic acid
derivatives and the p-nitrobenzyl ester with other
easily cleavable esters, suitable intermediates may
be prepared having the formula
N H H
3 \ ~
~ ,~ ~ CH2Y"
: 2
in which Y" is sulfonyloxy and R' is an easily cleavable
ester residue.
'
Preparation 3
~ Benzyl 7~-azido-3-bromomethyl- ~3-0-2-isocephem-
;~ 4-carboxylate
N3 \H H
n
o~ ~CH2Br
CH2c6H5
- 67 -
~' .
13~
To a solution of benzyl 7~-azido-3-hydroxy-
methyl- ~3-0-2-isocephem-4-carboxylate in benzene
is added about an e~uimolar amount of pyridine and
a slight molar excess of phosphorus tribromide.
There is produced the 3-bromomethyl title product.
The benzyl 7~-azido-3-hydroxymethyl- ~3-0-2-
isocephem-4-carboxylate used above may be prepared
by the method disclosed in Preparation 2 by substi-
tuting thebenzyl ester of the diiodide intermediate
for the p-nitrobenzyl ester used therein7
Other suitable 7/3-azido-3-halomethyl esters may
be prepared by the above method by appropriate vari-
ation of the ester acetoacetate starting material and
the halogenating agent.
: Preparation 4
Benzyl 7~-phenoxyacetamido-3-methyl-sulfonyloxy-
methyl- ~3-0-2-isocephem-4-carboxylate
C6H50CH2CO~H~
o \ ~ \ CH20502CH3
C02CH2C6H5
- 68 -
.
3~
Benzyl 7~-azido-3-methylsulfonyl-oxymethyl- .
~3-0-2-isocephem-4-carboxylate ~Preparation 2)
is dissolved in methylene chloride and cooled to
0C. Two equivalents of triethylamine are added
and, while stirring and cooling, H2S gas is passed
through the solution until it is saturated. The
solution is allowed to come to room temperature
and concentrated to give benzyl 7~-amino-3-methyl-
sulfonyl-oxymethyl- ~3-0-2-isocephem-4-carboxylate.
The ester is reacted with equimolar amounts
of phenoxyacetic acid and EEDQ in methylene
chloride to give the title product.
Other N-acylated compounds of the formula
H H
R-NH~
. I I
, o CH2oso2cH3
.
. may be prepared by substituting for the phenoxyacetic
~cid used above another desired acylating agent,
preferably one which will produce a starting material
in which R is one of the acyl groups mentioned as being
preferred in connection with the novel end-products of
formula II.
- 69 -
~ 3
Prep~r~tion 5
l-carboxymethyl-1~2,3,,4-tetrazole-5-thiol ~nd its
ti-soc~um salt
I
N N
Il 11
HS C\ /N
CH2COOH
A) Recrystallization of l-methYl-5-mercaptotetrazole
Procedure:
1. One hundred and ten grams of l-methyl-5-
mercaptotetrazole is slurried in 350 ml. of bo~ling
chloroform. A near solution is obtained.
2. The hot solut~on (50-60) is rapidly filtered
-; by vacuum through a heated ~uchner fu~nel (11 cm. SS
No. 604 paper containing 1/4 to 113 inch of packed
filter aid ("Supercel"). The filter pad is washed 7
with 50 ml. of 50-60 C. chloroform which is added to
the filtrate.
3. The filtrate is cooled to approximately 0-6 C.
ant kept at 0-6 C. for 2 hours. The crystals whlch
have formed are collected by filtration at 0-6 C. and
washcd with 60 ml. of 0-6 C. chloroform which is ~dded
to the filtrate. The crystals (cut A) are air drieA at
37-45 C. for 18 hours.
* Trademark.
-79-
4. The filtrate i8 concentrsted on the
rotary v8cuum evflporator t60~ C. bath) to ~pproxi-
mately one-half volume. This glurry is coolet to
0-6 C. end kept ~t 0-6 C, for 2 hours. The
cry~tals are c~llected by filtration at 0-6 C.,
washed wirh 40 ml. of 0-6 C. chloroform which is
edded to the filtrate. The crystals (cut B) a~e
air dried at 37-45 C. for 18 hours. Crystal cuts
A and B are compos~ted to give an approximate 65%
we~ght yield.
5. The filtrate of cut B, Step 4 may be
reworked tw~ce as described in Step 4 to o~tain
an additional 15% secovery.
B) Preparation of l-carboxymethYl-1,2,3,4-tetrazole-
5-thiol and its di-sodium salt
1. ~lve hundred ml. o~ subst2ntlally dry and
pure tetrahydroruran ln a 2-llter ~ neck flask with
~tlrrer ls cooled ln a salt-acetone-lce bath to
approximately -10 C. Dry nitrogen gas i9 blown
on the llquld surrace.
2. ~lve hundred ml. of 15.06~ (1.6 N) b~tyl
lithlum ln hexane (Foote Mineral Co.) ls added
over a ten minute perlod under dry nitrogen and
~tlrring to the tetrahydrofuran. me near
solutlon is cooled to -5 to -10 C.
-71-
~ 3 5
3. Fort~ ælx and rOur tenths gram (46.4 g,)
Or l ~ethyl-5-mercaptotetrazole (recrystallized
a~ QboYe) ls dlssolved ln 200 ml. Or substantlally
pure and dry tetra~ydro~uran. qhe solutlon 18
~lltered lr cloudy and then cooled to 5 to 10 C.
4. The cooled solution o~ step ~ 18 added
over 10 minu~e~ wlth stlrring and under dry nit~o-
gen to the butyl llthium solution. The temperature
should be maintalned at -5 C. to ~10 C. maxlmu~.
Preclpltates may form.
5. Ihe mixture 1~ stlrred under dry nltrogen
and 0 C. to ~10 C. for one halr hour.
6, Anhydrous carbon dioxlde gas ls bubbled
through at a rapld rate and wlth rapld stirrlng
ror 15-30 minutes at approxlmately amblent
temperature (0 to 10 C.) to no higher than
~20 C.
7, m e white prec~pitate whlch rorms ls
8ultab~y collected by rlltratlon in an area Or
low humidlty. The preclpltate ls wa~hed with about
75 ml. of tetrahydroruran.
8. The preclpltate ls dlssolved in 250 ml. ~ -
of water (pH 8.5-9.5). A second layer of tetra-
hydrofuran may be present. Thls may be removed
~n the Yacuum rotary evaporator (50 C. bath~, i
,
-72-
813S
9. The a~eous solutlon is adJusted to
pH 1.6-2.0 with concentrated hydrochlorlc acid.
10. me acld aqueou~ solutlon ls extracted
twice wlth 250 ml. portlons Or ethyl acetate.
Each 250 ml. ethyl acetate extract ls back
extracted wlth 100 ml. portions Or water. m e
water extracts are dlscarded. The ethyl acetate
extracts (rree o~ any water layer) are rlltered
and compo~lted.
11. m e comblned e~yl acetate extracts are
concentrated to dryness on the vacuum rotary
evaporator (60 C. bath),
12. m e cry~tals ln the flask are bolled with
~00 ml. Or chloroform for about 2 mlnutes. The
hot slurry (50-60 C.) ~s vacuum ~iltered through
a heated ~uchner ~mnel (11 cm-SS~604 paper), m e
crystals are washed with about 75 ml. o~ 50 C.
chloroform. The crystal~ are alr dried at room
temperature rOr about 3 hour~ and then made about
100-200 me~h,
13. The 100-200 mesh crystals are treated
with bolllng chloro~orm exactly as described ln
step 12 (the hot chloroform removes ~ost o~ the
unreacted l-methyl-5-mercaptotetrazole). Yield:
approxlmately 45 to 50 gram~ Or crystalline 1-
car~oxymethyl-1,2,3,4-tet~azole-5-thiol. These crystalc
~ J~ ~ 3~
uay contaln 0.02 to O.O5 moles Or l-methyl-5-
aercaptotetrazole .
1~ . me crystals o~ fitep 1~ are slurrled
~lth 250 ml~ Or ethyl ether at room temperature
~or ~-5 m~nutes. ~he mlxture ls filtered. The
lnsoluble~ (O.5-5%~ may be a contaml~atlng
symmetrical mercaptotetrazole ~etone o~ t~e
~ollowlng tentatlve structure:
N - ~ O N ~ N
~C ~N CH2 - C - CH2 - N ~ ~N
SH SH
CAUTIO~: Thls compound EXPLODES at approximately
205-210 C.
15. Ihe ether flltrate o~ 8tep 14 1~
evaporated to dryness on the vacuum rotary
evaporator (5O C. bath). Approximately ~2 to
48 grams Or crystalllne l-carboxymethyl-1,2,3.4-tetra-
zole-5-thiol containing approximately 0.01-0.05 mole
Or l-methyl-5-mercaptotetrazole 16 recovered.
16. The crystals are dicsolved ln 420 ml.
absolute ethanol (approxlmately lOO mg./ml.).
he solutlon ls warmed to 50-60 C.
17, TQ the hot ~olutlon Or step 16, 310 ml.
Or a 41% sodlum 2-ethylhexanoate (SEH) solutlon
ln i~opropanol ls added wlth very rap~d stlrr~ng
over a lO m~nute period. A crystalllne precipltate
ror~s. The ~.lxture ls ~lurried at 50-60 C, for
20 minutes.
74
:
J~135
~ 8 . me mixtuFe 1~ riltered hot (50-60 C.)
through a heated Buchner ~unnel ~11 cm-SS-No. 60
paper). m e crystals are washed wlth 75 ml. o~
50 C. ethanol.
19. ffl e ethanol damp crystal~ of ætep 18
- are slurried in 200-~00 ml. o~ ethanol. The
~lurry 15 passed through a 200 mesh screen. lhe
slurry ls heated to 50-60 C. ror 5 minutes wi~h
rapld ~lrring (unreacted di-~odlum l-methyl-5-
mercaptotetrazole is very so~uble ~n hot ethanol).
20. qhe crystals are collected at 50-60 C.
on a 11 cm-SS No. 604 paper ln a heated Buchner
~unnel. The crystals are washed wlth 75-100 ml.
ethanol and vacuum dried at 50-60 C. for
24-48 hours. yield: 40-48 grams Or dl-sodium
l-carboxymethyl-1,2,3,4-tetrazole-5-thiol (free of 1-
methyl-~-mercaptotetrazole as observed by h~).
-75-
.
1~ 13~ i
l-CarboxYethy1-1,2.3.4-tetrazole-5-~hiol
.
N - N
11 11
HS ~ /N
(CH2) 2-C02H
R) 2-CarboethoxyethYlisocYanate
~ -alanine ethyl ester hydrochloride (93.6 g.),
triethylamine (123.5g) and methylene chloride ~400 ml)
were mixed together and cooled to -10 C. Carbon
disulfide (46.S g) dissolved in 150 ml. of chloroform
was added to the above solution during a two-hour
period while keeping the temperature at about -10 C.
~fter the addition was complete, the temperature was
allowed to warm to 10 C. for a~out 10 minutes. The
501ut~0n was again cooled to 10 C. and 66.3 g of
etllyl chlorof~rmate in 60 ml of chloroform was added
dropwise over a 40-minute period with stirring. The
temperature was ~llowed to rise to room temperature
for 30 min~tes and again cooled to 0C. an additional
,.,
61.6 9 of triethylamine was ~dded a~ 0 C. and then
the 601ut~0n was stirred at room temperature for 3
hours.
~ he mixture was treated with water and the organlc
phase collected, washed w~th 2 x 250 ml of 2N HCl, then
2 x 250 ml of NaHC03, then 2 x 250 ml of water, The
osgan~c phase was dried over Na2S04 and the solvent
7~
1 ~ 81 3 ~ '
removed in vacuo to produce 93.7 ~ ~f ~n oil found
to be the desired product. The IR and NMR spectra
were consistent with the structure.
B) l-Carbox~ethyltetrazol-5-thio'
&odium azlde (29.7 g) was dissol~gd ~n 400 ml
~f water and heated to 60 C in a nitrogen ~tmosphere.
2-Car~oethoxyethylisocyanate (46.9 g) dissolved in
~0 ml of Skellysolve B (essentially n-hexane) wa
adde~ to t~e heated sodium azide ~olution. The
~olut~on was stirred for about 150 minutes at about
70-72 C., then cooled to 30 C. ~n an ice bath.
50~ sodium hydroxide solution was added until the pH
was 12. The mix~ure was heated for forty minutes at
70 C. and cooled to 15 C. in an ice ~ath~ The pH
was adjusted to 2 using conc~ Hcl and then extracted
wlth ethyl acetate t4 x 150 ml). The ethyl acetate
extracts were washed with water, then dried over
80dium sulfate. The sol~ent was evaporated 'n vacuo
~nd the product was collected as crystals from
methylene chloride to yield 19.5 g of ti~le product.
Substitution in the procedure for the pre-
paration of 1-carboxyethyltetrazol-5-thiol for
the ~-alanine ethyl ester used therein of ~n
equimolar quantity of an appropriately substituted
~mino acid ester of 3 ~o 4 carbon atoms produces the
correspond~ng l-carboxy tC3-C4 alkyl)tetrazol-5-thiol,
e.g., 1-carboxypropyltetr~zol-5-thiol and l-car~oxy-
bu~yltetrszol5-thiol.
* Trademark.
77
i~l`Q~.;3~
E X A M P L E_S
Example 1
; .
p-Nitrobenzyl 7~-azido-3~ methyltetrazol_5-
ylthiomethyl)- ~3-0-2-isocephem-4-carboxylate
N N
I I O N N
o,),_ ~fLC~2S~
C02CH2 ~)_ N2 ~ ~:
~ ` '
"
To a solution of p-nitrobenzyl 7~-azido-3-
methylsulfonyloxymethyl- ~3-0-2-isocephem-4-carboxYlate
(4.53 9., 10 mmole) and triethylamine (1.4 ml., 10 mmole)
in 90 ml. of methylene chloride was added 10 mmole of
l-methyltetrazole thiol. The solution was stirred at
24C. for 16 hours, then washed with 5% HCl and water
(100 ml. each), dried and the solvent evaporated in vacuo
to give the product as a yellow oil. Upon recrystalli-
zation from ethyl acetate, the title product was obtained
in 7~% yield; m.p. 150-152C. The NMR was consistent
with the proposed structure.
'
. .
78
7 ~ _
Example 2
p-Nitrobenzyl 7B-azido-3-(2-methyl-1,3,4-thiadiazol-
~-ylthlome~hyl)- ~3-0-2-isocepnem- ~
~,./\o ~ ,
o.~ CN2 5~5 1
co 2 cH 2~ N0 2
The procedure of Example 1 is repeated except that
the l-methyltetrazole thiol used therein is replaced
:
by 10 mmole of 2-methylthiadiazole thiol. The title
product was isolated as a yellow oil in 95~ yield.
The NMR and IR spectra of the product were in agreement
with the proposed structure.
Example 3
p-Nitrobenzyl-7~-azido-3-~2-methyl-1,3,4-oxadiazol-
5-ylthiomethyI)- ~3-0-2-isocephem-4-carboxylate
O ~ C 2 ~ O
C02CH2~ N2
,
_ 79 _
313~
A solution of p-nitrobenzyl 7~-azido-3-methylsUl-
fonyloxymethyl- ~3-0-2-isocephem-4-carboxylate (1.36 g.,
3.0 mmoles) and triethylamine (0.38 ml., 3.0 mmoles)
in 50 ml. of dichloromethane was treated with 2-methyl-
1,3,4-oxadiaæole-5-thiol (0.35 g., 3.0 mmoles). The
reaction mixture was stirred at room temperature for
48 hours and then washed with 10% HCl, water and brine.
The solution was dried over sodium sulfate and evaporated
in vacuo giving crude product. The crude product was
purified by dry-column chromatography on silica gel
- (60 g., Activity III) eluting with 15% ethyl acetate
in ether. There was obtained 1.0 g. of pure title product
in 70% yield. The NMR spectrum was in agreement with
;~ the proposed structure. ~ ;
Example 4
If the general procedures of Examples 1-3 are
repeated using equimolar weights of 1,2,3-triazole-5-
thiol, l-carboxymethyl-1,2,3,4-tetrazole-5-thiol and 1-
carboxyethyl-1,2,3,4-tetrazole-5-thiol, respectively,
in place of the thiols used therein, there are produced
p-nitrobenzyl 7~-azido-3-(1,2,3-triazol-5-ylthiomethyl)-
A3 o 2 isocephem 4 carboxylzte, p-nitrobenzyl 7~-azido-
~` ,
-- 80 --
13S
3-(1-carboxymethyl-1,2,3,4-tetrazol-5-ylthiomethyl)-
3-0-2-isocephem-4-carboxylate and p-nitrobenzyl
7~-azido-3-(l-carboxyethyl-l,2,3,4-tetrazol-5-
ylthiomethyl)- ~3-0-2-isocephem-4-carboxylate,
respectively.
Example 5
p-Nitrobenzyl 7~-amino-3-(1-methyltetrazol-S-
ylthiomethyl)- ~3-0-2-isocephem-4-carboxylate
H2 HCIH3
O N N
N ~ CH2-S J ~ l
CO2CH2 ~ - N
,
Hydrogen sulfide was slowly passed into a solution
of the 7~-azido ester of Example 1 (340 mg., 0.72 mmole)
and triethYlamine (77 mg., 0.76 mmole) in 10 ml. of
methylene chloride for 50 min. One ml. of 10% HCl
~, was added and the mixture concentrated in vacuo. The
` residue was mixed with 25 ml. methylene chloride and
; washed with 25 ml. each of 10% NaHCO3 and dilute NaCl.
The methylene chloride solution was dried, treated with
charcoal, filtered and evaporated ln vacuo. The re-
sulting residue was mixed with methylene chloride, the
sulfur filtered off and the solvent evaporated ln vacuo
to give the title product, 276 mg. ( 86% yield), as a
.81
yellow tar. The IR and NMR spectra were consistent
with the proposed structure.
Repeating the above procedure but using the benzyl
7~-azido ester instead of the p-nitrobenzyl ester gave
benzyl 7~-amino-3-(1-methyltetrazol-5-ylthiomethyl)-
~3-0-2-isocephem-4-carboxylate.
Example 6
p-Nitrobenzyl 7B-amino-3-(2-methyl-1,3,4-thiadiazol-
5-ylthiomethyl)- ~3~0-2-isocephem-4-carboxylate
H N H H
2 ``" r ~ oL C~~S ~ C~
c02CH2 ~r NO2
The procedure of Example 5 is repeated with the
p-nitrobenzyl 7~-azido-3~ methyltetrazol-5-ylthio-
methyl)- ~3-0-2-isocephem-4-carboxylate used therein
replaced by an equimolar weight of p-nitrobenzyl 7,3-
azido-3-(2-methyl-1,3,4-thiadiazol-5-ylthiomethyl)-
~3-0-2-isocephem-4-carboxylate. The title product
is produced as a yellow solid in 85% yield. The IR
and NMR spectra of the product were consistent with
the proposed structure.
' .
Example 7
p-Nitrobenzyl 7~-amino-3-(2-methyl-1,3,4-oxadiazol-
5-ylthiomethyl)- ~3-0-2-isocephem-4-carboxylate
- 82 -
~8135
H2N H _ ~ N N
~ ~ C~2-S ~ ~ c~3
C02CH2~ N2
Hydrogen sulfide was bubbled through a solution
of the 7B-azido ester of Example 3 (0.99 g., 2.1 mmole)
and trithylamine (0.32 ml., 2.3 mmole) in 50 ml. of
dichloromethane for 1 min. and the mixture was then
stirred for an additional 45 min. Nitrogen was then
bubbled through to remove excess H2S and the solvent
was evaporated in vacuo to leave a residue of title
product.
, .
Example 8
Pivaloyloxymethyl 7~-amino-3-(1-methyltetrazol-
5-ylthiomethyl)- ~3-0-2-isocephem-4-carboxylate
The title compound is produced according to
the method of Example 2 of U.R. Specification
1,229,453 by replacing the 7-aminocephalosporanic
acid used therein by 7~-amino-3-(1-methyltetrazol-
5-ylthiomethyl)- ~3-0-2-isocephem-4-carboxylic acid.
:`
'.~'
'~,
- 83 -
13~
The respective acetoxymethvl; methoxymethyl,
acetonyl and phenacyl esters of 7~-amino-3-(1-
nethyltetrazol-5-ylthiomethyl)- ~3-0-2-isocephem-
4-carboxylic acid are prepared by substituting in
the method above for the chloromethyl pivalate used
therein an equimolar weight of chloromethyl acetate,
chloromethyl methyl ether, chloroacetone and phenacyl
bromide, respectively. .
Example 9
p Nitrobenzyl 7~-(2-thienylacetamido)-3~ methyl-
tetrazol-5-ylthiomethyl)- ~3-0-2-isocephem-4-carboxylate
~, .
~ CH2CONS `O CNH3 N
C52-5 --( ¦
C2 2 ~ NO2
A solution of 2.0 mmole each of p-nitrobenzyl
7~-amino-3-(1-methyltetrazol-5-ylthiomethyl)- ~3-0-
2-isocephem -4-carboxylate, 2-thienyl acetic acid and
- 84 -
EEDQ in 20 ml. of methylene chloride was stirred at
24C. for 2 hours. The resulting solution was washed
with 5% NaHCO3, 10% HCl (2x) and dilute NaCl (20 ml.
each), dried and the solvent evaporated in vacuo
to give the product as a yellow oil. The product was
crystallized from methylene chloride/ether to give
the title product in 53% yield; m.p. 165-168 (dec.).
The IR and NMR of the product were consistent with
the proposed structure.
Example 10
p-Nitrobenzyl 7~-(2-thienylacetamido)-3-(2-methyl-
1,3,4-thiadiazol-5-ylthiomethyl)- ~3-0-2-isocephem-
4-carboxylate
s ~Lc~2s1\, C~3
C02CH2 ~ N02
The procedure of Example 9 was repeated with
the 7~-amino ester used therein rePlaced by
- 85 -
p-nitrobenzyl 78-amino-3-(2-methyl-1,3,4-thiadiazol-
5-ylthiomethyl)- ~3-0-2-isocephem-4-carboxylate.
The product was absorbed onto 6 g. of silica gel and
placed onto a 24 g. column of silica gel. The
column was eluted with methylene chloride followed
by ethyl acetate. The major fraction was obtained
as a yellow foam in 55% yield. Upon recrystalliza-
tion from methylene chloride/ether, the title
product was obtained; m.p. 127-129C. ~he IR and
NMR spectra were in agreement with the proposed
structure.
. ~ .
Anal. Calc'd. for C24H21N5O6S3: C, 49.05; H, 3.60;
N, 11.92; S, 16.37.
Found: C, 47.42; H, 3.54;
N, 11.87; S, 16.20.
Example 11
p-Nitrobenzyl 7~-phenoxyacetamido-3-(1-methyltetrazol-
S-ylthiomethyl)- ~3-0-2-isocephem-4-carboxylate
:~ .
. ~,
C6H OCH2CONH H H
~C~2_s~ ! :
CO2CH2 ~ 2
; A solution of p-nitrobenzyl 7~-amino-3-(1- -
methyltetrazol-5-ylthiomethyl)- ~3-9-2-isocephem-
4-carboxylate (270 mg.). EEDQ (150 mg.) and
phenoxyactic acid (92 mg.) (0.60 mmole each) in
6 ml. of methylene chloride was maintained at 24C.
for 2 hours. This solution was diluted to 15 ml.
with methylene chloride, washed with 5% NaHCO3
(25 ml.), 10% HCl (2 X 25 ml.) and saturated MaCl
(25 ml.), an~ then dried and absorbed onto 1.7 g. of
silica gel (grade III). The silica gel was placed on
a chromatographic column containing 3.4 g. of silica
gel (grade III). The column was eluted with ether
changing to ether-methylene chloride tl:l) after
fraction 4 (all fractions were 5 ml.). TLC of fractions
8-17 showed them to contain one compound (Pf=0.11 on
silica gel eluted with ether). These fractions were
combined and evaporated in vacuo to gi~e the title
product as a yellow tar, 236 mg. (67% yield). The
.
.
-~
, ~ .
~ - 87 -
.
11~8~;~
i
IR and NMR spectra of the product were consistent
with the proposed structure.
Example 12
Benzyl 7~-phenoxyacetamido-3-(1-methyltetrazol-5-
ylthiomethyl~- ~ 3-0-2-isocephem-4-carboxylate
The procedure of Example 11 was repeated except
that the p-nitrobenzyl startin~ material was replaced
by the correspondinq benzyl ester. The title product
was obtained upon recrystallization from methylene
chloride-ether (1:1); m.p. 86-88C. U.V.~ meaxH 284
; (=10,400). The NMR and IR spectra of the product
- were consistent with the proposed structure.
Example 13
p-Nitrobenzyl 7B-(2-thienylacetamido)-3-(2-methyl-
1,3,4-oxadiazol-5-yl)- ~3-0-2-isocephem-4-carboxylate
; I I H H
CH2CONH~ CH2 sl~ o J~ CH3
C2 CH2 ~3 N02
: .,
- 88 -
.~ - .
11~ 35
The p-nitrobenzyl ester residue of Example 7
was dissolved in 50 ml. of dichloromethane and
treated with 2-thienylacetic acid (0,31 g., 2.2
mmoles) followed by EEDQ (0.54 g., 2t2 mmoles).
The mixture was stirred at room temperature for 16
hours and then washed with water, 1% NaHCO3, water,
10% HCl, water and brine. It was then ~ried over
sodium sulfate and evaporated in vacuo. ~he residue
was purified by dry-column chromatography on silica
gel (50 g., activity III) eluting with 25~ ethyl
acetate in ether to give 0.61 g. of pure title
product in 50~ yield. The NMR of the product was
in agreement with the proposed structure.
Example 14
7B- (2-Thienylacetamido)-3-(1-methyltetrazol-5-
ylthiomethyl)- ~3-0-2-isocephem-4-carboxylic acid
:
The p-nitrobenzyl ester from Example 5 (1.00
mmole) in 50 ml. of ethyl acetate and 25 ml. of n-butanol
to which 10 ml. of 0.1 M HCl and S00 mg. of 20% palladium
hydroxide-on-diatomaceous-earth had been added was
hydrogenated on a Parr apparatus at 24C and 50 p.s.i.
..:
- B9 -
11~ 35
for 3 hours. The catalyst was filtered off and the
~olution extracted with 1% NaHCO3 (containing some
MaCl) (3 X 25 ml.). The aqueous extract was cooled
to 0C., acidified to pH 1 with 10~ HCl and saturated
with NaCl. The acidified aqueous was extracted with
methylene chloride (4 X 50 ml.). The methylene
chloride was concentrated in vacuo to 100 ml. and
after standing, the title product was collected as
a colorless solid in 18.5% yield; m.p. 182-184
,....~
(dec.). U.V. ~mhax 280 (~=8500). The IR spectrum
was consistent with the proposed structure.
';
Anal. Calc d- for C16H16N6 5 2
N, 19.26.
Found: C, 44.10, H, 3.71;
N, 19.07.
A sample of the title product ~called BC-L70)
; after solution in water and dilution with Nutrient
~roth was found to exhibit the following Minimum
Inhibitory Concentrations (M.I.C.) in meg./ml. versus
the indicated microorganisms as determined by over-
night incubation at 37C. by tube dilution.
-- 90 --
M.I.C. in mc~./ml.
. Or~an~sm BC-L70 ~eph~lexin CePhalothin
D~ pneumonlae A9585 .016 ..25 .13
~5% 6erum*
Str. pyosenes A9604 .016 .25 . .06
~57. serum* -~
S. eureus Smith~ A9537 .13 1 .13
S. sureus Smith~ A9537 1 1 .5
~50% 8erum
S. sure~s BX1633-2 A9606 .5 2 .25
et 10 dil'n
S. aureys BX1633-2 A9606 2 4 .5
et 10 ' dil'n
S. ~ureus meth.- A15097 1 8
resist.; at 10-3
dil'n
Sal. enteritidist A9531 .03 4 .5 ~ :
E. coli Juhl~ A15119 1 8 16
E. col~$ A9675 16 8 63
. pneumoniae$ A9977 .25 4 4
K. pneumoniae$ A15130 8 16 16
Pr. mirabil~s$ A9900 .13 4
Pr. morgani~ A15153 32 125 125
P8. aeru~inosa~ A9843A 125 125 125
Ser. marcescens~ A20019 125 125 125
Ent. cloacae A9656 125 125 125
Ent. cloacae A9657 1 4 4
Ent. cloacae A9659 125 125 125
* 50% Nutr~ent Broth - 45% Antibiotic Assay Broth
at 10 4 d~lue~on.
--91-
Example 15
7~-(2-Thienylacetamido)-3-(2-methyl-1,3,4-thiadiazol-
5-ylthiomethyl)- ~3-0-2-isocephem-4-carboxylic acid
The procedure of Example 14 was repeated up to
the point of preparing the acidified aqueous extract
with the p-nitrobenzyl ester starting material re-
placed by p-nitrobenzyl 7B-(2-thienylacetamido)-3-
(2-methyl-1,3,4-thiadiazol-5-ylthiomethyl)- ~3-o-2-
isocephem-4-carboxylate. The acidified aqueous was
extracted with tetrahydrofuran/ethyl acetate (9:1)
(2 X 50 ml.).
The organic extract was washed with saturated
NaC1 (50 ml.), dried, treated with charcoal and the
solvent evaporated ln vacuo to give a brown oil.
The oil was crystallized from ethyl acetate/ether
to give the title product in 39% yield; m.p. 100-
105C. (dec.). U.V. ~mHaF/H2O 276 (= 12,500). The
IR and NMR were consistent with the proposed structure.
Anal. Calc~d. for cl7Hl6N4o5s3-l/2 CH3CO2C2H5: ~
C, 45.96; H, 4.06; N, 11.29; S, 19.37.
; Found:
C, 45.69; H, 3.97; N, 11.30; S, 19.63.
. .
- 92 -
135
M.I.C. data for the product (called BC-L71) is
shown in the following table.
M.I.C. in mc~./ml.
~ BC-JJ71 Cephalexin ~ephalothin
D. pneumoniae A9585 .016 5 .06
~5% ~erum* .008 .5 .06
Str. pyogenes A9604 .016 .25 .06
~5% serum* .008 .13 .06
S. ~ureus Smith i A9537 .25 1 .13
.06 ~5 .06
S. aureus Smith ~ A9537 2 2 .5
~50X serum 2 2 .5
S. aureus BX1633-2 A9606 .5 2 .25
at 10-3 dil'n .13 1 .13
S aureus BX1633-2 A9606 4 4 ,5
a~ 10-2 dil'n 2 2 .2S
S. aureus meth.- A15097 4 16
res~st.; at 10-3 2 16
dil'n
Sal. enteritidis ~ A9531 06 2 13
E. coli Juhl $ A15119 8 8 8
E. coli ~ A9675 322 16 32
K. pneumoniae i A9977 ,5 4
K. pneumoniae ~ A15130 32 166 16
Pr. mirabilis ~ A9900 1 8
Pr. morganii ~ A15153 132 125 125
Ps. aeruginosa ~ A9843A 125 125 125
Ser. mascescens ~ A20019 125 125 12$
Ent. cloacae A9656 125 125 125
Ent. cloacae A9657 4 4 4
Ent. cloacae A9659 125 12255 1225
* 50% Nutrient Broth - 45% Antibiotic Ass~y Broth
at 10 4 ~ilution.
Q~13~
Example 16
7~-~2-Thienylacetamido)-3-(2-methyl-1~3,4-oxadiazol-
5-ylthiomethyl)- ~3-0-2-isocephem-4-carboxylic acid
The p-nitrobenzyl ester of Example 13 (0.60 g.,
0.15 mmole) was dissolved in 125 ml. of ethyl
acetate and 40 ml. of n-butanol. The solution was
treated with 1.05 ml. of IN HCl (1.05 mmole) and
hydrogenated over 0.60 g. of 20% palladium hydroxide
on carbon for 3 1/2 hours at 50 p.s.i.g. The mixture
was then filtered, washing well with ethyl acetate,
and evaporated in vacuo. The residue was slurried
with ether and extracted with 1% NaHCO3 (2 X 50 ml.)
followed by 25 ml. of water. The aqueous extracts were
acidified in the cold with 10% HCl and extracted with
ethyl acetate (3 X 30 ml.). The solution was dried
over sodium sulfate and evaporated ln vacuo. The
solid residue was slurried with ether, filtered
and dried giving 69 mg. (15% yield) of the title
product; m.p. 170-173. U.V.~ max = 278 nm (F = 12,453).
20Anal. Calc'd. for C17H16N4O5S2: ,
N, 12.84.
~`` Found: C, 43.95; H, 3.57;
N, 11.35;
residue: 3.05.
M.I.C. data for the product (called BC-L76) is
shown in the following table.
- 94 -
L3
M,l.C. in mc~./ml.
~anism BC-L76 Ccphalexin Cephalothin
D. pneumoniae A9585 .016 .~5 .016
~5% ~erum*
Str. pyogenes A9604 .008 .13 .03
+5% ~erum*
S. aureus Smith ~ A9537 .06 .5 .06
S. aureus Smith ~ A9537 1 1 .25
~50% serum
S. aureus 8X1633-2 A9606 .5 2 .13
at 10-3 dil'n
S. aureu3 BX1633-2 A9606 .5 4 .13
at 10-2 dil'n
S. aureus meth.- A15097 2 32 2
resist.; at 10-3
dil~n
Sal. enteritidis ~ A9531 .5 2 .13
E. coli Juhl $ A15119 8 8 16
E. coli ~ A9675 63 16 63
K. pneumoniae i A9977 1 4 2
K. pneumoniae ~ A15130 63 8 32
Pr. m~rabilis ~ A9900 .5 4 .5
Pr. mor~anil ~ A15153 125 ~125 >125
Ps. ~eru~inosa $ A9843A ~125 >125 ~125
Ser. marcescens ~ A20019 >125 ~125 ~125
Ent. cloacae h9656 ~125 >125 ~125
Ent. cloacae A9657 16 , 4 8
Ent. cloacae A9659 ~125 ~125 ~125
* 50% Nutrient Broth - 45% Antibiotic Assay Broth
at 10 4 dilution.
-- 95 --
13:~
7~-phenoxyacetamido-3~ me~hyltetrazol-5-ylthio-
methyl~ 3-0-2-isocephem-4-carboxylic acid
A solution of the p-nitrobenzyl ester prepared
according to Example 11 (~3~ mg., 0.405 mmole) in
12 ml. of tetrshytrofuran (peroxide free) and 2.4
ml. of ethanol with 140 mg. of 10% palladium-on-
charcoal was hydrogenated (Parr Shaker) at 24
and 50 p.s.i. for 4 hours. The cstalyst was
filtered off and the solvent evaporatet in vacuo.
The residue was dissolved in 25 ml. of ethyl
acetate and washed with 25 ml. each of 10% HCl
and water. The acqueous layers were combined
and extracted with 10 ml. of ethyl acetate (which
was washed with 10 ml. of water). The com~ined
ethyl acetate was extracted with 1% NaHC03
96-
'
~ 35
(2 x 10 ml.). The combined bicarbonate extracts
were acidified with 10% HCl and extracted w~th
ethyl acet~te. The ethyl acetate extract was
wa~hed with water ~nd ~aturated NaCl nd then
drled and evaporated ~n vacuo to give 67 mg.
(37% yield) of crude title product. The product
was crystallized from chloroform to give title
product with m.p. 136-138 (dec.). U.V. ~ MaeH
276 (~=10,600).
Anal. Calc d. for C18 18 6 6
N, 18.83; S, 7.18.
Found: C, 46.70; H, 3.94
N, 17.74; S, 6.98;
Residue: 1.16.
.
M.l.C, data for the product (called ~C-L62)
is shown in the following table.
1~ 35i
.
M.I.C._ in mc~ . Iml .
Or~anism BC-L62 Cephalexin Cephalothin
D. pneumoniae A9585 .13 .13 .03
~SX ~erum*
Str. pyogenes A9604 .13 .06 .03
~5Z serum*
S. sureus Smith i A9537 .25 .25 .06
S. aureus Smith ~ A9537 1 1 .25
~50% ~erum
S. ~ureus BX1633-2 A9606 2 2 .13
at 10-3 dil'n
S. aureus BX1633-2 A9606 16 4 .25
at 10-2 dil'n
S. aureus meth.- A15097 4 16
reslst.; at 10-3
dil'n
Sal. enteritidis ~ A9531 .5 4 .13
E. coli Juhl $ A15119 4 8 16
E. coli ~ A9675 32 16 32
K. pneumoniae i ~9977 2 4
K. pneumoniae ~ A15130 32 16 32
Pr. mirabilis ~ A9900 2 4 .5
Pr. mor~anii ~ A15153 63 125 >125
Ps. aeruginosa ~ A9843A ~125 ~125 ~125
Ser. ~arcescens ~ A20019 ~125 ~125 >125
Ent. cloacae A9656 ~125 ~125 ~125
Ent. closcae A9657 32 4 4
Ent. cloacae A9659 125 >125 ~125
* 50% Nutrient Broth - 4S% Antibiotlc Assay Broth
~t 10 4 dilution.
--98--
11~ 3~ ~ .
Example 18
am~no-a-phenvlacetamido]-3~ methyltetrazol- -
5-YlthiomethY~ 3-O-?-lsocephem-4-carboxyllc ~cid
(h~drochloride salt)
H-CONH ~ 0 IH3
2 o~ ~CH2-S ~
C02H HCl N--N
A mixture contain~ng p-nitrobenzyl 7~-amino-3-
(l-methyltetrazol-5-ylthiomethyl)- A3-0-2-isocephem-
4-carboxylate (2.8 g., 6.25 mm~le), EEDQ (1.55 g.,
6.25 mmole) and D(-) -azidophenyl acetic acid ~1.11 g.,
6.25 mmole) in 200 ml. methylene chloride was kept at
room temperature (protected from moi~ture by ~ calcium
chloride drying tube) for 16 hours; It wa6 then washed
successively with 10% HCl, water, 5% NaHC03 and brine,
dried (Na2S04) ~nd evapor~ted in vacuo to dryness leaving
a yellow amorphous solid. This was ~uspended in ether
~nd filtered to glve 3.67 g. of a solid whlch was
identlfied by IR and ~MR a6 p-nitrobenzyl 7~ azido-
~-phenylacetamido]-3-(1-methylt~trazol-5-ylthiomethyl)-
3-0-2-isocephem-4~c~rboxylate; m.p. 132-134C.
'"
~ .
_99_
11~8135
An~l. Calc'd- for C25H22N107S C~ 49-S0; H~ 3~65;
N, 23.09.
Found: C, 49.47; H, 3.64;
N, 22.86.
A mixture consistlng of the above ester (0.40 g.,
0.65 mmole) snd 20% Pd(OH)2 on Celite t0.50 g.) in 50
ml. ethyl ~cetate, 50 ml. n-butanol And 6.5 ml. of .
O.lN HCl was 6haken on a Parr apparatus under 60 p.~
of hydrogen for 6 hours. The solid was filtered off
on a Celite pad which was washed well with absolute
ethsnol. The combined filtrate6 were evaporated in
VACUO at 40C. to leave ~ yellow flmorphous ~olid. This
was triturated with ether and collected by filtratio~
to give 0.21 g. solid; m.p. 110-120 (deco~p,), U.V.
= 274, ~ maX ~7550. The IR and NMR ~pectra of
the product were in agreement with.the proposed structure.
M.I.C. data for the product (called BC-L78) ls shown
in the following table, Purity of the sample was estin~ted
to be bout 25%.
.~ .
.
-100- . '
35 ~
: i
M.l.C. ln mc~.~ml.
Or~anism BC-L78 ~ Cephalothir
D. pneumoniae A9585 0.25 0.25 .03
~SZ serum~
Str. pyogenes A9604 0.13 0.13 .03
~5X serum*
S. ~ure~s Smith ~ A9537 2 0.5 .06
S. aureus Smith ~ A9537 8 1 0.5
~50% serum
S. sureus BX1633-2 A9606 8 2 0.25
~t 10-3 dil'n
S. ~ureus BX1633-2 A9606 63 8 0.5
at 10-2 dil~n
S. sureus meth.- A15097 (37) 63 32
resist.; at 10-3 (28)>125 li5 63
- dil 'n
Sal. en~erltidis ~ A9531 0.5 2 0.25
E. coli Juhl ~ A15119 2 8 16 : :
E. coli ~ A9675 16 16 63
K. pneumoniae ~ A9977 1 4
K. pneumoniae ~ A1513~ 8 16 16
~r. mirabilis ~ A990n 1 4
Pr. ~organ~i ~ A15153 32 ~125 ~125
Ps. aeruginosa i A9843A >125 >125 ?125
Ser. ~arcescens ~ A20019 >125 >125 ~125
Ent. cloacae A9656 ~12~ 125 ~125
Ent. cloacae A9657 4 4 4
Ent. cloacae A9659 16 ~125 ~125
* 50% Nutrient Broth - 45% Antib~otic Assay Broth
nt 10 4 dilution.
- 101 --
-
w~
Exsmple 19
. Repe~ting the general N-acylatlon procedures of
Ex~mple6 9, 1~ 11, 12, 13 , or 18 to react the following
~cylating agents with 7~-amino-3~ methyltetrazol-5-
ylthiomeehyl)-~\3-0-2-i~ocephem-4-carboxylic acid (or
~n ester or salt thereof), the following products ~re
obt~ined after removal of sny functional blocking groups.
.,,,~ ' .
AcYlatin~ ARent Product
~ixed anhydride of 7~-(2-Aminomethylphenyl-
potassium 2-(1-carbo- scetamido)-3-(1-methyltetrazol-
methoxypropen-2-ylamino- 5-ylthiomethyl)-~3-0-2-
methyl)phenyl~ceta~e with lsocephem-4-carboxylic acid
isobutyl chloroformate
~-benzoylureidophenyl- 7~ -Benzoylureidophenyl-
~cetic acid ~cetamido)-3-(1-methyltetrazol-
S-ylthiomethyl)- ~3-o-2-
. .
. lsocephem-4-carboxylic acid
, ~
2,6-dimethoxybenzoyl ~ ,6-Dimethoxybenzamido)-
; chloride . 3-(1-methyltetrazol-5-ylth~o-
. ~ethyl)- ~ -0-2-isocephcm-4-
carboxylic acid
~ .
, .
~ -10~- -
. - . . .
Acylatin~ A~ent Product
D-~nhydro-o-carboxy- 7~-(D--Hydroxyphenylacetemido)~
~ndelic acid 3~ methyltetrazol-5-ylth~o-
~ethyl)- A3_0_2_isocephem-4-
carboxylic acid
3 benzyl-1,2,4-oxadiazole- 7~-~N-(Phenylacetimidoyl)-
S-one-4-acetic ~cld sm$noacetamido~-3-(1-methylte-
trazol-5-ylthiomethyl)-~ -0-2-
~socephem-4-carboxylic ~c~ d
.
; ~aler~c acid 7~-Valeramido-3-(1-methylte-
trazol-5-ylthiomethyl)- ~3-o-2- ~:
- 160cephem-4-carboxylic ~cid
phenylace~ic acid 7B-Phenylacetamido-3-(1-
methyltetrezol-5-ylthiomethyl)-
A3 0 2 isocephem-4-carboxylic
acid
3-thienylacetyl chloride 7~-(3-Thienylacetamldo)-3-
(l-methyltetrazol-5-ylthio-
methyl)-~3-0-2-isocephem-4-
' carboxylic acid '
a-carboxybenzyl-phenyl- 7~ carboxY-~-Phenyl-
~cetic acid acetamido]-3-~1-methyltetrazol-
~: 5-ylthiomethyl)- ~3-0-2-isoccphem-
4-c~,rboxylic ~c~d
-103-
.
8~ 3~
~c~laeinr~ ~ent Product
~-hydroxyphenylacetic 7~-(o-Hydroxyphenylacet~mido)-
cld 3-(l-methyltetrszol-5-ylthio~ethyl~-
~3-0-~-isocephem-4-carboxyl$c
acid
cysnoacetlc acid 7~[cyanoacetamidol-3-(l-
methyltetrazol-5-ylthiomethyl)-
/\3-0-2-isocephem-4-carboxyllc
acid
~-cyanopropionic ~cid 7~-(a-cyanopropionamido)-3-
~l-methyltetrazol-5-ylthio-
methyl)-~3-0-2-isocephem-4-
c~rboxyl$c ~cid
~-(2H)-tetrazoleacetic 7~-[2-(2H)-tetrazolylacetsmido~-
ac$d 3-(1-methyltetr~zol-5-yl~hiomethyl)-
: ~3-0-2-isocephem-4-carboxylic acld
3-(o-chlorophenyl)-5- 7~-[3-(o-chlorophenyl)-5-
methyl-4-isoxazole- ~ethylisoxazol-4-ylcarboxamidol-
carboxylic scid chloride 3-(l-methyltetr~zol-5-ylthio-
methyl)- ~\3-0-2-$socephem-4-
csrboxylic acid
l-(lH)-tetrazolylacetyl 7~-ll-(lH)-tetrazolylacetamido]-
; chlor~de. 3-(l-meehyltetr~zol-5-ylthi
methyl)-~3-0-2-isocephem-4-
carboxylic acid
- 104 -
l~Q~3~ ~
EX~
~ ollowing the ~cyl~t~on ~ethots of the preceeding
examples and in particulsr those d~sclosed in U.S. ~atent
3,546,219, the compounds listed below sre prepared by
reacting 7~-a~lno-3-(1-methyltetrazol-5-ylthiomethyl)-
~3-0-2-isocephem-4-c~rboxylic acid (os an ester or salt
~heseof), wi~h the ~pproFriate scylsting agent.
RYCO-CO~ ~ fH3
N
o~N~
C02H N -
RY_
phenyl
p-acetamidophenyl
p-methoxyphe~yl
p-methylphenyl
2-methoxy-5-methyl~henyl
m-chlorophenyl
o-nitrophenyl
2,4-d~chlosophenyl
a-naphthyl
2-phenanthryl
p-aminophenyl
2-thienyl
p-~imethyl~minophenyl.
' A r
~ #13
x~mple 21
Follow~ng the acyls~on methods of ~he preceeding
cx~mples ~nd in particular those di~closed in U.Kc
Patents 1,296,081 and 1,294,541, ~he compounds l~sted
~elow are prepsred by re~cting 7~-a~ino-3-tl-~ethyltetrazol-
S-ylthiomethyl)-~3-0-2-isocephem-4-carboxylic ~cid or sn
e8ter or 8alt thereof with an 2cylating agent of the
formNl~
Ra -CR-COOH
X--C
H-N-C-R
Il
. X'
or 2 funct~onal equivalent thereof.
X C ~CH25~
H-N-C-R C02H N--N
X '
Ra X X' ~
phenyl 0 ~mino ~H2
2-Shienyl 0 i~ino ~H2
~; 3-thienyl 0 imino NH2
trophenyl 0 ~mino NH2
O ~mino NH2
~-aminophenyl
p-methylplleny~ 0 imino ~H2
~chlo~ophenyl 0 imino N~2
B~,
Ra X X' R~
p-methoxyphenyl O ~mino N~2
p-hydroxyphen~1 0 imino ~2
p-di~hylamlno- O ~mino NH2
phenyl
3,4-dimerhoxy- 0 ~mino N~2
phenyl
~-methoxyphenyl O i~o N~Z
p-ace~amidcphenyl O imino ~H~
m-hydroxyphen~-1 0 ;~;no ~2
3~5-dichl~ro-4- 0 Imino NH2
hyds~xyphenyl
3-chloro-4- 0 imino ~H2
: hydrox~phenyl
phenyl O 0 2-fury~
2-thienyl 0 ~ 2-f
3-~hienyl 0 ~ 2-fury~
phenyl O O phenyl
2-tbienyl 0 0 phenyl
phenyl 0 0 2-thieny~
p-chloroph~nyl O 0 2-uryl
p-hyd~oxyphen~l O 0 2-~ury
3-chloro-4- 0 0 2-fury~
hydrox~phenyl
3~5-dichloro-~- 0 0 2-furyl
hyd~oxyphenyl
: ~fl7
il~8~3
Ra X Xi R~
m-aminophenyl O 0 2-furyl
p-methylphenyl O 0 2-furyl
p-dimethylamino- O O ~-furyl
phenyl
p-me~hoxyphenyl O 0 2-furyl
~-hydroxyphenyl O 0 2-furyl
p-acetamidophenyl O 0 2-furyl
m-nitrophenyl O 0 2-furyl
phenyl O O CH3
2-thienyl 0 0 CH3
3-thienyl 0 0 CH3
phenyl O O -CH2-C6H5
phenyl O O
phenyl O O
phenyl O O
phenyl O ~l3C ~
N~o CH3
phenyl O O ~ ~
~O~CH3
phenyl O N~S~
ir~i8
~ 35
Ra X X ' Ri
phenyl O o
phenyl O O N~O~Lc6Hs
phenyl O O ,¢~
-CH2
phenyl S 0 2-furyl
2-thienyl S 0 2-fu~yl
3-thienyl S 0 2-furyl
p-hydroxyphenyl S O CH3
phenyl O imino phenyl
phenyl O imino 2-thienyl
phenyl O imino 2-fury
3-thienyl 0 imino phenyl
phenyl O imino l I
' , '
9-
1 ~ 81 3 5
Ex~mPle ?2
When 7~-amino-3-(1-methyltetrazol-5-ylthiomethyl)-
/\3-0-2-isocephe~-4-carboxylic acid or an ester or salt
thereof is acylated according to the procedures above
and ln particulsr those tisclosed in U.S. 3,692,779
with an acid chloride of the formula
RlTN - CH2COC 1
~,~ ~0~0
; there are produced the compounds listed below.
.
CH
R -~ -CH2CONH~ N--N
O ~ 2 ~
C02H N N
'
R~
benz)~l ;
-110-
813;~ii 1
~,S ; ~ ; ~,~ ;
N~ N3 ; ~
C6H5 ~1~3 ; 3 ~bL ; ~;
dichlor om~ thyl ;
n-propyl;
cyclopentyl;
., ,
cyclohe~cyl;
p-chlosobenzyl;
~ phenyl
: 2-thie~yl;
3-thienyl. ~ :~
... .
.
~8~13
EX8mD1 e 2 3
When the 7-acylamido-3~tl-m~thyltetrazol-5-
ylthlo~ethyl)- ~3-0-2-isocephem-4-car~oxylic acid
compounds of Example 28 ~se hydrogenated as by the
process of U,S, Pate~t 3,692,~79, these are produced
the compounds listed bel~wO
Ri- o ~~~C~2CO~Y.~ l H3
o~ ~ C~12 ~
co2~ N- N
where Ri is as de~ ned in Example 22.
Example 24
When 7~-amino-3-(1-~ethyltetrazol-5-ylthio~ethyl)-
~3-0-~-isocephe~-4-car~oxylic acid o~ an ester or salt
thereof is acylated according to the procedures of the
above examples (an~ in p2rticulas the prccedures disclosed
in U,S. Patent 3,646,024) with an acid chloride of the
formula
; R -CH-CO-C1
R It
'
.ere sre prsduced the compounds listed below.
~ 81~5 '~
Ra-CH-CONH CH3
R~ N NH ~ 1_N
o o~N~CH~~S_~
Ra Ri
phenyl phenyl
phenyl 2-thienyl
phenyl 2-furyl
2-thienyl phenyl
3-thienyl 2-$uryl
p-hydroxyphenyl phenyl
. . .
ExamPle 25 ~ :
When 7B-amino-3-(1-methyltetrazol-5-ylthiomethyl)-
~3-0-2-isocephem-4-car~oxylic acld or an ester or salt
theseof is acylated ~ccording to the procedures sbove
~nd in particular sccoTding to the methods of U.S.
Patent 3,778,436 with ~n acylat~ng agent of the formula
R -CH-COOH
.,, S-C-R
i 11
O
or a functlonal equivalent thereof, there are produced
the compounds l$sted below
-113-
1 1~ 8 ~ 3 5
!
l~-CH-CON}I~-- 2
R R~
.
3,4-d~ethoxyphenyl ~thyl
2-thienyl
p-methylphenyl
2,4-dichloro?henyl ethyl
S-methyl-3-phenyl- ethyl
~80~azol-4-yl
2-thienyl ethyl
2-furyl
2-furyl
phenyl
phenyl
1,4-cyclohexadien- ~ethyl
l-yl
-il4-
1~0~L3 ~ -
~!i ,
Preparation of 7~-t~-~2-AminomethYl ~ Ey~hexadien~
cetamido~-3-(1-methyltetrazol-5-ylthiometh~l~- L~3 0-2-
isoce~hem-4-carboxylic acid
~CH2CONH ~f o
~ C02H N - N
,.~ ~ ' .
A. a-(2-Amlnomethvl-1~4-~yclohexadienyl~aceti~ acid
A solution of 16.5 g. ~0.1 mole) of o-aminomethyl- '`!
phenylacetic acld in 1.5 1 of llqu~d ammonia (which had
been treated with 50 ~. of Li to remove a trace of :~
moisture) was slowly diluted with 500 ml. of dry t-BuOH.
To the solution was addet in sm~ll portions 3.4 g. (0.5g,-
: atom) of Li over a period of 4 hours and ~he mixture
i .~"
i was stirred for 16 hours at room temperature removing
:
.. the liquid ammonia in a hood and finally evapora~ed to
dryness.below 40 C. The residue was dlssolved in 500 ml.
of water and ~he solution was chromatographed on a column
,
--115-
of IR-120 (H~, 700 ml.) res~n ~nd el~ted with lX
NH40H solution. N~nhydr~n positive frac~,lons of
the eluate were combined and evaporated to dryne~.
The residue wa ~ashed with four 50 ml. portions of
hot acetone ~nd recrystallized from 500 ml. of ethanol-
water (1:1) to give 11.2 8. (677.) of a-(2-amin~methyl-
1,4-cyclohexad~enyl)acetic acld as colorless needles.
M.p. 183C.
IR: ~ nu~ 1630, 1520, 1380, 1356 cm 1.
~MR ~D2O ~ R2C3 2.72 (4H, ~, H2C_ ), 3-01
(2H, s, CH2CO), 3.20 (2H, 5, CH2-N), 5.78 (2H, s, H_ C=).
Anal. Calcd. for CgH13N02 C, 64.65; H, 7.84;
N, 8.38.
Found: C, 64.77; ~, 8.06;
N, 8.44.
8. a-[2-(t-Butoxycarbonvlaminom thvl~-1,4-cyclohexadienyl]-
scetic acid
'
To a ~tirred solution of 8.0 g. (0.048 mole) of
a-(2-aminomethyl-1,4-cyclohexadienyl)acetic acid and
3.8 g. (.096 ~ole) of NaOH in 150 ml. of water wa.c added
a solution of 10.3 g. (0.072 mole) of t-butoxycarbonylazide
ln 80 ml. of THF and the mixture was stirred.for 18 hours
-1~6-
~08~3~
~t r~om temper~ture. The THF was removed u~der
reduced pressure and the residual solution was
~a~hed with eeher (2 x 100 ~1.), acidifiet with
6 N HCl ~nd extracted w~th ether (3 x 100 ml.). The
combined extraces were washed with ~ater (2 x 100
~1.) and ~ saturatet NaCl solution ~lO0 ml.),
dr~ed with Na2S04 and evaporated to dryness. The
oily residue was triturated with n-hexane to give
10.5 g. (82%) of colorless p~wder ~elting at 113C.
IR: ~/ ma~ 3370~ 1715, 1640, 1530, 1280, 1160 cm 1.
NMR: ~ CpC13 1.45 (9H, s, t-Bu-~), 2.73 (4H, s, H2C ~C C)~
3.16 (2H, s, CH2C0), 3.76 (2H, d, 6Hz, CH2N)
4.90 (lH, ~, NH), 5.66 (2H, s, H> C=), 10.6 (lH,
br-s, C00~,
Anal. Calcd. for Cl4H21N04: , ; .
Found: C, 63.13; H, 8.21; N, 5.26.
C. 7B-lG-t-ButoxYcarbonYlaminomethy~ _clohexadienvl)-
acetamido~-3-(l-methyltetrazol-5-ylthiomethyl)-~3-0-
2-isoce~hem-4-c2rboxylic a _d
'
To a stlrred solution of equimolar amounts of ~-12-
(t-butoxycarbonylaminomethyl)-1,4-cyclohexadienyl)acetic
scid and 2,4-din~trophenol in ethyl acetate is added an
equ~molar emount of N,N'-d~cyclohexylcarbodiimide. The
~eac~ion mixture ls stir~et a~ room tempersture for
3 hours. She ~eparated d~cyclohexylurea is filtered
off. ~he filtrate is evaporated to dryness to give
the ~ctivsted ester which is dissolved in eetrahydr
~uran. To this solut~on is added a solution of 7B-
~mino-3-(1-~ethyltetrazol-5-ylthio~ethyl)- ~3-o-2-
~ocephe~-4-carboxylic acid and triethylamine in
~pproxim~tely a 1:2 molar pro?ortion, respectively,
relative to the a-[2-(t-butoxycarbonylam~nomethyl)-
1,4-cyclohexadienyl]scetic acid. The mixture is stir~ed
~t room temperature for several kours and concentrated
in V2C~O. The concent_ate is washed with ether, ac~difiet
with dilute mineral acit and estracted with ethyl acetate.
The extracts are washed with water and sa~urated NaCl
solution and dried to gi~e the title product.
D. 7B~ (2-Amino~eth~l-l,~-cvclohexadionvl~acetamito~-
3-(l-m2ehvltetrazol-S-YlthicmethYl~- G3-0-2-isoce~he~-
4-carbox~ acid
A solution of 7B-~c-(2-t-bu~oxyca-Donyl2mino~.ethyl-
1,4-cyclohaxadienyl)aceta~ido]-3-(1-methyltetrazol-5-
ylthiomethyl)-~3-0-2-isoce?hem-4-ca bo~ylic acit ~n
trifluoroacetic acid is stirred at 0C. for one ho~r.
1~C1 8~3S `~
~o the solution ~s ~tded dry ethes un~il a prec~pitate
forms. The precipitate is collected by filtration,
~u~pended ~n water Jn~ ad~usted ~o pH6 to give the title
produc~.
ExamPle 27
7-~ ~a-(2-Am-inomeehv~ -cyclohe~xenvl~acetamidol-3
tetrazol-5-Ylthio~ethvl~ 3-0-2-isoce~he~-~-c~r~oxvlle
acid
~ CH2-s~
A. ~?-(N-t-~tox~ar~o~Ylaminsmethvl~ c~clohe,~en-
-acetic acid
A solution of ~-[2-(t-butoxycarbonylaminome~hyl)-
1,4-cyclohexadienyl3-acetic acid (1.33 g., 5 mmoles)
ln 3~. a~mon~ hydroxi~e (10 ml.) was hydrogenated at
40 psi with pallad_us2 o~ cha-eoal (lOZ, O.~ g.). A
th~oret~cal amount of hydrogen w~s taken up in 3 hours.
~he ca~alyst was removed and the filtrate uas acidified
~o pH 2 w~th dil. ~Cl a~t extracted with ethyl ~cetate
11~
3L~
(2 x SO ml.). The co~bined extracts were washed with
~ater (20 ml.), dr~ed with Na2S04 ~nd evfipor~ted ~nder
redueed pressure to afford an oil (1.34 g.) which ~ol~di-
f~ed on ~tanding for several days. Recrystall~zation from
n-hexsne - ethyl ~cetate gave 1.2 g. t~tle product as
colorless pris~s melting st 118-119C.
IR ~ nu~ol 3450, 173~, 1660, 1510 cm
NMR:~ ppm 3 1.58 (9H, s, t-butyl-~), 1.50 - 1.90
(4H, m, -CH2-), 1.90 - 2.20 t4H, m, llylic
methylene-H), 3.18 (2H, s, C~2-CO), 3.78
(2H, d, 6 Hz, CH2-N), 5.00 (lH, br-s, NH),
8.98 (lH, br-s, COOH).
: Anal. Calcd. for C14H23N04: C, 62-43; H, 8-61;
N, 5.20
Found: C, 62.12; H, 8.77;
N, 5.37.
B. 7~-[~-(2-t-~utoxycarbonYlaminomethv-l-cYclohexenvl)
scetamido~-3-(1-methvltetrazol-5-ylthiomethvl)-~3-
0-2-isoceDhem-4-carbDxylic acid
: To a stlrred solution of equimolar amounts of
2-(N-t-butoxycarbonylaminomethyl)-l-cyclohexen-l-yl]
acetic acid snt 2,4-dinitrophenol ~n erhyl acetate is
-120-
.
1~8i~
added an equimolar a~ount of NIN'-d~cyclohexylcarbo-
diimide. The reactlon mixture is fitirret for 1 hour
at room temperature and the precipitated d~cyelohexylurea
1~ filtered o~f. The filtr2te is cooled to 5-C. and
poured into a cold ~olution of 7~-amino-3-(1-methyl-
te~razol-5-ylthiomethyl)- ~3-0-2-isocephem-4-carboxylic
acid snd excess triethylamine in 50% ~queous T~F. The
mixture is stisred overnight at room temperature and
washed with ether. The aqueous layer is acidif~ed
with tilute HCl to precipitate the title produet.
,
C. 7~ Aminomethyl-l-cyclohexenyl~acetamido~-3-
(l-methyltetrazol-5-vlthiomethYl~- A3-o-2-
isocePhem-4-~arboxYlic acid
' ' .
A solution of 7~-[c-(2-t-butoxycarbonylaminomethyl-
l-cyclohexenyl)acetamido]-3-(1-methyltetrazolo5-ylthio-
~ethyl)- ~3-0-2-isocephem-4-c~r~oxylic acid in trifluoro-
~cetic ac$d is stirret at 0C. for 1.5 hours. The mix-
ture is diluted wi~h ethe. to separate the trifluoro-
ecet~te salt which is dissolved in water and neutralized
to give the title p~oduct.
Exam~le 28
~'
When the p-nitrobenzyl 7~-amino-3-(1-methyltetrazol-
5-ylthiomethyl)-~3-0-2-isocephem-4-carboxyl~te of
Ex~mple 15 ls replaced by sn equimolar weight of p-
~21
1 ~ 8 i 3 5
nltrobenzyl 7~-smino-3-(1,2,3~triszol-5-ylthio-
~ethyl)-~3-0-2-~socep~em-4-c~rboxylete, p-niero-
benzyl 7~-amino-3-(1-carboxymethyl-1,2,3,4-tetrazol-
S-ylth~omethyl)-~3-0-2-isocephem-4-carboxylate ~nd
p-nitrobenzyl 7~-amino-3-(l^carboxyethyl-1,2,3~4-
tetrazol-5-ylthi~methyl)-~\3-0-2-isocephem-4-carboxylate,
respectively, there are produced after catalytic hydro-
genation as in the procedure of Example 20 7~
thienylacetamido)-3-~1,2,3-triszol-5-ylthiomethyl)-
~3-0-2-isocephem-4-car~oxylic acid, 7~-(2-thienylacet-
~mido)-3-(1-carboxymethyl-1,2,3,4-tetrazol-5-ylthio-
methyl)-~3-0-2-isocephem-4-carboxylic acid snd 7B-
(2-thienylacetamido)-3-(1-carboxyethyl-1,2,3,4-
tetrazol-5-ylthiomet~.yl)-~3-0-2-isocephem-4-carboxylic
acid, respectively.
Example 29
When the 7B-amino-3-(1-methyltetrazol-5-ylthiomethyl)-
~3-0-2-isocephem-4-carboxylic acid (or ester or salt
thereof) in the procedures of Examples ll , 12 , 18 and
19-27. is replaced by an equimolar amount of 7~-amino-
3-(1,2,3-triazol-5-ylthiomethyl)- ~3-0-2-isocephem~4-
carboxylic acid, 7~-amino-3-(2~methyl-1,3,4-thiadiazol-
--l22-
~0813~ i
5-ylthiomethyl)- ~3-0-2-isocephem-4-carboxylic arid, 7~-
amino-3-(2-methyl-1,3,4-oxadiazol-5-ylthiomethyl)- ~3-0-2
isocephem-4-carboxylic acid, 7~-amino-3-(1-carboxymethyl-
tetrazol-5-ylthiomethyl)- ~3-0--2-isocephem-4-carboxylic
acid or 7B-amino-3-(1-carboxyethyltetrazol-5-ylthiomethyl)-
~3-0-2-isocephem-4-carboxylic acid (or an ester or fialt
thereof, any reactive functional groups other than the 7- ~
amino group being suitably protected if necessary), respec- ~ -
tively, there are produced (after any necessary deblocking
of functional protecting groups) the corresponding 7~-acyl-
amino carboxylic acids of each of the above-named nuclei.
Example 30
.;
When the d-amino products of any of the aforementioned
Examples pertaining thereto are reacted with acetone accord-
ing to the procedure of U.S. Patent 3,303,193, there are ob-
tained the corresponding 0-2-isocephem derivatives of the
formula
Ra-CIH c----n CH ~
HN N CH CH O
C C N
H3C / ~ H3 O I CH2-S-Z
COOH
where Ra is as defined in the above-mentioned examples
and Z represents the appropriate alkyl, aryl, aralkyl
or heterocyclic group of the selected nucleus, or
pharmaceutically acceptable salts thereof.
, ~
- 123 -
8i3
.
Ex~mPle 3l
7~-Phenox~acetamido-3~ methYltetrazol-5-Ylthio- -
meth~ 3-0-2-~socePhem-4-carboxYlic acid
(~lternate process)
A mixture o~ benzyl 7~-phenoxyacetamido-3-
meehylsulfonyloxyme~hyl- ~3-0-2-lsocephem-4-
c~rboxylate tO.22 g., 0.43 mmole; preparet accordi~g
to preparation 4 ~bove), triethylam~ne (0.07 ml.,
0.5 mmole) and l-methyltetrazole thiol ~O.OS g., -
0.5 mmole) ~n methylene chloride ~50 ml.) was stisred
at room temperat~re for 16 hours, washed with lOZ HCl
~20 ml.) and brine ~2 x 50 ~l.), dried and co~centr~ted
to give a semi-solid (0.24 g.) which was purlf~ed by
column chromatography to afford 0.15 g. of pure benzyl
7~-phenoxyacetamido-3-~l-methyltetrazol-5-ylthiomethyl)-
~3-0-2-isocephem-4-carboxylate ~s a wh~te sem~-solid.
The product was shown by IR snd NMR to be identical
with the product of Example 18
Anal. Calc'd. for C25H24N6065
N, l5.66.
~ound: C~ S5.89; H, 4.55;
N, 15.34.
The benzyl ester ls subjected to catalytlc
hydrogenation followin~ the procedure of Example -~
23 to give the title product.
4-
