CA1072543A

CA1072543A - Cephalosporin compounds

Info

Publication number: CA1072543A
Application number: CA245,853A
Authority: CA
Inventors: David A. Berges
Original assignee: Smith Kline and French Canada Ltd
Current assignee: GlaxoSmithKline Inc
Priority date: 1975-02-27
Filing date: 1976-02-16
Publication date: 1980-02-26
Also published as: DE2607681A1; FR2302098A1; JPS51110595A; GB1544703A; IL49066A0; BE838760A; AU1140676A; IE42961L; FR2302098B1; ZA76451B; IE42961B1; AU499565B2; IL49066A; NL7601935A; IT1055451B; LU74427A1

Abstract

ABSTRACT
The compounds of this invention are cephalosporins having a thienyl- or tetrazolylacetamido substituent at the 7-position and a carboxy or carbamoyl substituted triazolyl- or tetrazolylthiomethyl group at the 3-position of the cephem nucleus. The compounds have antibacterial activity.

Description

Sf~3 This invention relates to a new series of cephalo-sporin compounds which have antibacterial activity when ad-ministered parenterally. In particular, the compounds of this invention are characterized by having a carboxy or car-bamoyl substituted triazolyl or tetrazolylthiomethyl group ; at the 3-position of the cephem nucleus.
The compounds of this invention are represented by the following structural formula:

'' 10 1 S

:~ ~ N ~
. O I CH2SHet .
~ COOH

: Formula I
in which:

Rl is thienyl or tetrazolyl; and . Het is selected from the group consisting of:

^~ \
N_N ~N~

(CHR2)n-CoR3 H (CHR )n-CoR3 ~CHR2) -CoR3 : m , . in which each individual R2 is hydrogen or lower alkyl, n is zero to ten, m is one to ten and R3 is hydroxy, amino, lower alkylamino or di(lower)alkylamino, or a non-toxic pharmaceutically acceptable salt thereof.
`- As used herein, the term "lower alkyl" refers to ~ 30 groups having from one to our carbon atoms, especially :: methyl and ethyl.
, ~ -2-:' , ~ ~ zr~

Preferred compounds of this invention are represent-ed by Formula I in which Het is tetrazolyl substitu~ed with -(CHR )mCoR3 where R is hydrogen, m is one to -ten and R3 is hydroxy, amino, lower alkylamino or ditlower)alkylamino.
Advantageous compounds of this invention are represented by Formula I in which Het is tetrazolyl sub-stituted with -(CHR2) CoR3 where R2 is hydrogen, m is one to five and R3 is hydroxy or amino.
Particularly preferred are the compounds 7-(2-thienylacetamido)~3-(1-carboxymethyltetrazol-5-ylthiomethyl)-3-cephem 4-carboxylic acid, 7-(2-thienylacetamido)-3-[1-(2-carbamoylethyl)tetrazol-5-ylthiomethyl]-3-cephem-4-car-. boxylic acid, 7-(1-tetrazolylacetamido)-3-(1-carboxymethyl-tetrazol-5-ylthiomethyl)-3-cephem-~-carboxylic acid, 7-(2-thienylacetamido)-3 [1-(2-carboxyethyl)tetrazol-5-ylthiomethyl]-3-cephem-4-carboxylic acid, and 7-(1-tetra-zolylacetamido?-3-[1-(2-carboxyethyl)tetrazol-5-ylthio-methyl]-3-cephem-4-carboxylic acid.
Cephalosporin derivatives having a thienylacetamido or tetrazolylacetamido group at the 7-position are well documented in the prior art.
Cephalosporins of the present invention, substituted by a substituted S-heterocyclicthiomethyl group at the 3-position of the cephem nucleus, are encompassed by a generic formula in Netherlands Patent 6916151 where Het includes triazolyl and tetrazolyl optionally substituted with, inter alia, carboxy, carbalkoxy, alkoxyalkylaminocarbonyl and ` dialkylaminoalkylaminocarbonyl. This formula also contains . an optionally substituted heterocyclic acetamido group at the 7-position, among which are included tetrazolylace-tamido and thienylacetamido. Specifically, only an ester of a ~'7'~

carboxylic acid substituted tetrazolylthiomethyl cephalosporin ; or the present invention is disclosed.
Japanese Patent 7205550 contains a generic formula for cephalosporin compounds of this invention where Het in-cludes triazolyl and tetrazolyl substituted with -(CH2)nR3, where n is 0 to 3 and R3 includes alkoxycarbonyl, carboxy, N-alkoxyalkylcarbamoyl and dialkylamino and also having a 7-thienylacetamido or 7-tetrazolylacetamido group. Exem-plified are a 7-tetrazolylacetamido-3-(ester and acid sub-10- stituted thiadiazolyl)thiomethyl cephalosporin.
Recently issued U.S. Patent 3,819,623 contains a generic formula for cephalosporins of this invention bearing a 7-heterocyclicacetamido or 7-heterocyclicthioalkylacetamido group and having in the 3-position, inter alia, tetrazolyl-thiomethyl substituted with carboxy, alkoxycarbonyl, carboxy-alkyl, alkoxycarbonylalkyl and dialkylaminoalkylaminocarbonyl-alkyl. Among the compounds disclosed are an ester of a com-pound of this invention, and a carboxylic acid substituted thiadiazolylthiomethyl analog.
The compounds of Formula I are prepared by acyla-tion of an appropriate 7-amino-3-substituted triazolyl- or tetrazolylthiomethyl cephalosporin nucleu~ of Formula II:

' NH2 S

0//~----~ N ~ CH2SHet COOR

Formula II
in which , . .
~ 30 ~let is selected from the group consisting of:

,,, ~

H

N N _ N N -N
and ~/ ~

tCHR )n-COR H (CHR )n-COR (CHR )m-COR

in which each individual R2 is hydrogen or lower alkyl, n is zero to ten, m is one to ten and R3 is hydroxy, amino, lower alkylamino or di(lower)-alkylamino; and R is hydrogen or a protecting ester group, with thienyl or tetrazolyl acetic acid followed by removal of the protective groups. The carboxylic acid group is activated by any of the standard methods such as conversion to the mixed anhydride, acid chloride, acid imidazolide or activated ester.
In addition, a reagent such as dicyclohexylcarbodiimide can ~ be used provided that the carboxyl group on the cephem nucleus ;~ is protected with an easily removable protecting group such as a benzhydryl, t-butyl, trichloroethyl~ benzyl, benzyloxy-methyl, p-nitrophenyl, p-methoxyphenyl, p-methoxybenzyl or p-nitrobenzyl ester.
Alternatively, the compounds of Formula I are pre-¦ pared by acylating 7-aminocephalosporanic acid with thienyl or tetrazolyl acetic acid and then displacing the 3-acetoxy group with the desired substituted triazole or tetrazole thiol.
The protective groups can be removed according to methods well known to the ar-t, such as with trifluoroacetic acid when t-bu-tyl protective groups are used. The resulting salt is converted to the free acid by means of a basic ion exchange resin such as polystyreneamine ion exchange resin (Amberlite ~ IR-45) or else by basification of an aqueous solu-tion of the salt.

_ 5 _ . .

~ .
~V'^~5~3 The thienyl and tetrazoyl acetic acid starting mate-: rials are known or are prepared by known methods. 7-Thienyl-acetamidocephalosporanic acid and 7-tetrazolylacetamidocephalo-sporanic acid are also known (U.S. 3,S16,997).
The 7-amino-3-substituted triazolyl- and tetra-zolylthiomethyl cephalosporin starting materials of Formula II are prepared from reaction of 7-aminocephalosporanic acid ` and a substituted triazole or tetrazole thiol.
The substitu-ted 1,2,3-triazole thiols no-t known ` 10 to the art are prepared by rearrangement of a correspondingly substituted amino thiadiazole according to the procedure - of Goerdeler and Gnad [Chem. Ber. 99:1618 (1966)], by con-.~
version of a suitably substituted hydroxy 1,2,3-triazole to the corresponding thiol by the method of Hoover and Day [J. Amer. Chem. Soc. 78:5832 (1956)] or by a reaction of an - acetylene carboxylic acid with a substituted azide and sub-sequent decarboxylation and thiation.
The substituted tetrazole thiols where R3 is hydroxy are prepared by reaction of an isothiocyanate, for example ethyl isothiocyanoacetate, or an N-alkyl dithiocar-bamate, such as methyl 2-carboxyethyldithiocarbamate, with an azide such as sodium azide. When R is amino, lower alkylamino or di(lower)alkylamino, the tetrazole thiols are prepared from the corresponding tetrazole thiols where R3 is hydroxy by standard methods for the preparation of amides from acids, for example, by reaction of a tetraæole thiol where R3 is hydroxy with l,l-carbonyldiimidazole and an amine of the formula N~R5R6 where R5 and R6 are each hydrogen or lower alkyl, or by conversion of the -tetrazole -thiol where ; 3 R is hydroxy to the corresponding acid chloride with sub-sequent reac-tion of the acid chloride with an amine (NHR5R ).
., The tetrazole thiols are also prepared by conversion of a sui-tably substituted hydroxy tetrazole to the corresponding thiol by the method of Hoover and Day [J. Amer. Chem. Soc.
78:5~32 (lg56)].
The compounds of this invention are capable of forming salts with, for example, the alkali metals such as sodium or potassium, the alkaline earth metals such as calcium or with the ammonium cation. These salts are prepared by standard methods using a wide variety of non-toxic pharmaceu-tically acceptable acids and bases known in the art and are also considered as objects of this invention.
It will be recognized that due to the potentially asymmetric carbon atom in the 3-heterocyclic side chain of Formula I, optical isomers may exist. Racemic or resolved products are obtained depending upon whether a racemic or resolved side chain acid is used as an acylating agent. The resolved side chain acids are readily obtained from the racemic compounds by resolution according to well known methods, including fractional crystallization of a salt formed with an optically active base. All of the isomers, including separated isomers and mixtures thereof, are included within the scope of this invention.
The compounds of Formula I have antibacterial acti-vity against both Gram-positive and Gram-negative organisrns.
They also exhibit high serum leve:Ls and serum half-life values.
Minimum inhibitory concentrations (MIC's) ranged from 0.2 to ~200 ~g./ml. in in vitro testing. These results are shown in Table 1 below for a number of compounds of Formula I. In vivo mouse protection data are given in Table 2. Compound names corresponding to numbers are given in Table 3.

5~3 . . . ~ .
~ ~ o o~9 o o oIn oo Lr~ o o o o n ~ oo ~ o ~, ~ ~, A A A
::

~D ~ ~ O O O ~ O O O ~ O~D 0 ~1 0 O O O O O
~1~I t~l ~ t`l A A

U~ ~~D O O ~ ~ ~ r~ O~9 ~O O ~ O
O O ~1 0 00 ~1 0 . A A A

~ ~ ~ 00 ~~9 ~ ~ ~ Ln ~
_ ~ ~OOO~OOO~O~O
O O 0 ~10 0 ~1 A A A
., ~ I
., ~1 1':1 H~ ~J O ~1 ~J O
:' ~ ~ ~ ~
~3 A A

. ~ ~ ~ ~ CO ~ ~ 0~ 0CO
. ........ o O O ~ ~ O ~~1 0 ~`IO O '10 ~1 0 .~ . ` ` ` ` ` ` ` ` A ` ` Ll-) ` O
. ~r ~ ~u~ ~9 ~~D CO ~~t' ., t~l .,................ O O O ~ ~ ~ ~ O OO O O O ~9 0 ., ~ O ~ O
N ~I

r--l CO r--l~r) r-l ~ CO~I r-l ~1 ~ O O O ~ D ~ O O O ~ In o ~ o . InIn O ~ O
A A
, . CO
,.
CO OO U~
~,~ Ll~) ~CO ~I
.' ~ O O ~ ~0~ ~1 . ~ o o 1--., ~ ~ ~ ~1 O ~r--I ~1 0 51 1~
.,`~a~ ~ ~ ~r--l ~_) m r--l ., ~) X ~; r-l ~1 ~ O O 1~ 0 1 O ~ U~ N~a E3 .~ P:l ~10 ~_1 ~ ~t) ~ m ~ h r-l O
~ r-l tl) a)~J4-1 r~ r~
S I hr~ l r-l O ~--1 0~ O
O O ra Oa) hn~
a) a) ~~i~rl ~~1 ~J O

JZ~'~3 ED in vlvo (mg./kg.) Compound E. coli 12140 ~leb. pneumo. 4200 , :, s.c. p.o. s.c. p.o.
\
1 21.. ~ - 3.9 ~
. 2 1.56 46 1.56 35 3 7.2 35 1.0 40 4 0.4 >50 1.8 >50 5 21.2 ---- 25 -~--6 1.12 >50 1.56 -~
7 -___ ____ .
; Compound Number Compound Name 1 7-(2-thienylacetamido)-3-(1-carboxy-methyltetrazol-5-ylthiomethyl)-3-cephem-4-carboxylic acid .~ 2 7-(2-thienylacetamido)-3-[1-(2-carbamoylethyl)tetrazol-5-ylthio ~: 20 methyl]-3-cephem-4-carboxylic acid 3 7-(2-thienylacetamido)-3-[1-(2-carboxyethyl)tetrazol-5-ylthiomethyl]-3-cephem-4-carboxylic acid 4 7-(1-tetrazolylacetamido)-3-(1 carboxymethyltetrazol-5-ylthiomethyl)-3-cephem-4-carboxylic acid 7~ tetrazolylacetamido)-3-(5-carboxy- -.
methyl-1,2,4-triazol-3-ylthiomethyl)-3-cephem-4-carboxylic acid : 30 6 7-(1-tetrazolylacetamido)-3-[1-(2-carboxyethyl)tetrazol-5-ylthiomethyl]-3-cephem-4-carboxylic acid 7 7-(2-thienylacetamido)-3-[1-(10-.:- carbamoyldecyl)tetrazol-5-ylthiomethyl]-: 3-cephem-4-carboxylic acid Pharmaceutical compositions having antibacterial activity which comprise a pharmaceutical carrier and a compound of Formula I and methods of treating or preventing .: bacterial infections by admi.nistering a compound of Formula I to an animal in a nontoxic amount sufficiènt to treat or prevent said infection are also objects of -this invention. The administration may be by parenteral in-jection such as subcutaneously, intramuscularly or intra- -venously. The injection of suitably prepared sterile solu-tions or suspensions containing an effective, nontoxic amount of the new cephalosporin compound is the preferred route of administration.
The compounds of Formula I are formulated and administered in the same manner as other cephalosporins in dosages of from 250 to 1000 mg. with the total daily dosage being from 1 to 6 g. The precise dosages are dependent upon the age and weight of the subject and on the infection being treated and can be determined by those skilled in the art - based on the data disclosed herein compared with that available to the art attained with known cephalosporins.
The following examples illustrate the invention, but are not to be construed as limiting the scope thereof.
- Temperatures are in degrees Centrigrade unless otherwise stated.

7-(1-Tetrazolylacetamido)-3-(5-carboxymethyl-1,2,4-triazol-3-ylthiomethyl)-3-cephem-4-carboxylic acid To a solution of 1.05 g. (12.5 mmol.) of sodium bicarbonate in 30 ml. of water was added 1.36 g. (7.5 mmol.) of 3~carboxymethyl-1,2,4-triaæol-5-thiol sodium salt, pre-pared by addition of one equivalent of a sodium hydroxide solution to a solution of 3-carboxymethyl-1,2,4--triazol-5-thiol in aqueous ethanol, and 1.91 g. (5.0 mmol~) of 7-(l-tetrazolylacetamido)cephalosporanic acid. The reaction mixture was heated at 67.5 for four hours while maintaining ~ 5~

the pH a-t 7.6 by addition of glacial acetic acid. The mixture was cooled to 20, acid was added to bring the pH to 3.5 and the resulting solution was lyophilized and the residue trit-urated with ethanol. The product was dissolved in 15 ml. of absolute methanol and 0.196N solution of sodium methoxide in methanol was added until ca. pH 6Ø Ethyl acetate was then added to the solution and the precipitate was collected and dried in vacuo to give 7-(1-tetrazolylacetamido)-3-(5-carboxy-methyl-1,2,4-triazol-3-ylthiomethyl)-3-cephem-4-carboxylic acid disodium salt.
H N O S 2 Na 3 5 H O
Calculated: 31.98% C; 3.74gO H; 21.18% N; 10.54% S
Found: 32.53% C; 3.14% H; 20.72% N; 10.55% S
7-(1-Tetrazolylacetamido)-3-(5-carboxymethyl-1,2,4-triazol-3-ylthiomethyl)-3-cephem-4-carboxylic acid ; disodium salt is dissolved in a minimum amount of water to which ethyl acetate is added. While stirring, 3N hydro chloric acid is added until the solution is acidified to pH 2.5. The layers are separated, the aqueous phase is extracted with ethyl acetate and the combined extracts are washed with water, dried (MgSO4) and evaporated to dryness to give the title compound.

7-(2-Thienylacetamido)-3-(5-carboxymethyl-1,2,4-triazol-3-ylthiomethyl)-3-cephem-4-carboxylic acid When an equivalent amount of 7-(2-thienylacetamido)-cephalosporanic acid is substituted in the procedure of Example ; 1 for 7-(1-tetrazolylacetamido)-cephalosporanic acid~ the title compound is obtained.
The title compound may be converted to the corres-ponding disodium salt by addition of sodium methoxide solution as described in Example 1 ' ~

7~ Tetrazolylacetamido)-3-(4-carboxy-1,2,3-triazol-5-ylthiomethyl)-3-cephem-4-carboxylic acid _ To a solution of 4.6 g. (0.20 mol.) of sodium and 250 ml. of absolute ethanol was added under a nitrogen atmo-sphere 8.65 g. (0.05 mol.) of 5-amino-4-carbethoxy-1,2,3-thiadiazole. The reaction mixture was stirred one hour, 5 ml. of water was added and the mixture was refluxed for 12 hours. After cooling, the mixture was filtered and the solid product was washed with ethanol and ether and dried to give 4-carboxy-1,2,3-triazole-5-thiol trisodium salt.
Acidification of an aqueous solution of 4-carboxy-1,2,3-triazole-5-thiol trisodium salt gives 4-carboxy-1,2,3-triazole-5-thiol. The corresponding mono-sodium salt is prepared as described in the procedure of Example 1.
Substitution of an equivalent amount of 4-- carboxy-1,2,3-triazole-5-thiol sodium salt in the procedure of Example 1 for 3-carboxymethyl-1,2,4-triazole-5-thiol sodium salt gives the ti-tle compound.
` 20 EXAMPLE 4 7-(1-Tetrazolylacetamido)-3-(4-carbamoyl-1,2,3-triazol-5-ylthiomethyl)-3-cephem-4-carboxylic acid . . . _ Reaction of 7-(1-tetrazolylacetamido)cephalo-sporanic acid, 4-carbamoyl-1,2,3-triazole-5-thiol sodium salt, prepared from 4-carbamoyl-1,2,3-triazole-5-thiol as described above, and sodium bicarbonate according to the procedure of Example 1 gives the title compound.

7~ Tetrazolylacetamido)-3-[4-(2-carboxyethyl)-1,2,3-~ 30 triazol-5-yl-thiomethyl]-3-cephem-4-carboxylic acid - To a solu-tion of 1.725 g. (75 mmol.) of sodium - in 50 ml. of absolute ethanol was added 6.0 g. (45 mmol.) of benzylazide and 12.3 g. (50 mmol.) of diethyl 2-(carbo-methoxyethyl)malonate. The reaction mixture was refluxed for 12 hours, then cooled and evaporated to dryness. Water (100 ml.) was added and the mixture was heated while main-taining the pH at 12. The aqueous mixture was extracted with ethyl acetate and the aqueous phase was acidified with 3N
hydrochloric acid and extracted with ethyl acetate. The ethyl acetate solution was evaporated to dryness to give l-benzyl-4-(2-carboxyethyl)-5-hydroxy-1,2,3-triazole.
l-Benzyl-4-(2-carboxyethyl)-5-hydroxy-1,2,3-triazole (5.0 g., 0.02 mol~) was suspended in 300 ml. of ethanol. An-hydrous hydrogen chloride gas was bubbled in-to the suspension for 30 minutes and the mixture was then heated on a steam bath for 20 minutes. The solution was cooled and evaporated ! to dryness to give a residue which was dissolved in 300 ml.
of ethyl acetate. The ethyl acetate solution was washed with water, dried (~gSO4) and evaporated to dryness. Trituration with hexane containing a little acetone gave l-benzyl-4-(2-carbethoxyethyl)-5-hydroxy-1,2,3-triazole.
A solution of 0.63 g. (2.3 mmol.) of 1-benzyl-4-(2-carbethoxyethyl)-5-hydroxy-1,2,3-triazole in 0.70 g.
(3.5 mmol.) of phenylphosphonic dichloride was heated under ~` a nitrogen atmosphere a-t 160 for 1.5 hours. Water (2 ml.) and 2 ml. of chloroform were added -to the reaction mixture and it was stirred at 5 while 6 ml. of 5~ aqueous sodium bicarbonate was added. The mixture was extracted with chloroform and the extract was dried (Na2SO4) and evaporated to dryness to give 1-benzyl-4~(2-carbethoxyethyl)-5-chloro-1,2,3-triazole.
A solution of 0.54 g. (0.01 mol.) of sodium ::

' .

me-thoxide in 50 ml. of absolute ethanol is saturated with hydrogen sulfide. l-Benzyl-4-(2-carbethoxyethyl)-5-chloro-1,2,3-triazole (2.9 g., 0.01 mol.) is added and the reaction mixture is refluxed for 24 hours. The cooled mixture is evaporated to dryness to give l-benzyl-4-(2-carboxyethyl)-1,2,3-triazole-5-thiol sodium salt. Acidification of an aqueous solution of the triazole thiol salt as previously described gives l-benzyl-4-(2-carboxyethyl)-1,2,3-triazole-5-thiol.
1-Benzyl-4-(2-carboxyethyl)-1,2,3-triazole-5-thiol (1.1 g., 4 mmol.) is suspended in 50 ml. of anhydrous liquid ammonia and sodium is added until a permanent blue color results. The reac-tion mixture is allowed -to stir for 40 minutes then allowed to warm to ambient tempera-ture while the ammonia evaporates. The residue is triturated with ether and the solid formed is collected and dissolved in water.
The aqueous solution is acidified and extracted with ethyl acetate. The extract is dried (MgSO4) and evaporated to dryness to give 4-(2-carboxyethyl)-1,2,3-triazol-5-thiol.
The corresponding sodium salt is prepared as described in Example 1.
Substitution of 4-(2-carboxyethyl)-1,2,3-triazole-5-thiol sodium salt in the procedure of Example 1 for 3-carboxymethyl-1,2,4-triazole-5-thiol sodium salt gives the title compound.

: Use of equivalent amounts of diethyl 2-carbethoxy-methylmalonate and diethyl 2-(carbethoxypropyl)malonate/
respectively, in the procedure of Example 5 in place of di-ethyl 2 (carbomethoxyethyl)malonate, followed by the steps ` of ester hydrolysis, chlorination, sulfide displacement and ''' ~ - 14 -.

debenzylation described therein gives 4-carboxymethyl-1,2,3-triazole-5-thiol and 4-(3-carboxypropyl)-1,2,3-triazole-5-thiol. The corresponding sodium salts are prepared as des-cribed in the procedure of Example 1.
Use of 4-carboxymethyl-1,2,3-triazole-5-thiol sodium salt and 4-(3-carboxypropyl)-1,2,3-triazole-5-thiol sodium salt in the procedure of Example 1 in place of 3-carboxymethyl-1,2,4-triazole-5-thiol sodium salt gives, respectively, 7-(l-tetrazolylacetamido)-3-(4-carboxymethyl-1,2,3-triazole-5-ylthiomethyl)-3-cephem-4-carboxylic acid and 7-(1-tetra-zolylacetamido)-3-[4-(3-carboxypropyl)-1,2,3-triazol-5-ylthiomethyl]-3-cephem-4-carboxylic acid.

To a solution of 13.8 g. (0.6 mol.) of sodium in absolute ethanol is added a solution of 96 g. (0.6 mol.) of diethylmalonate in 500 ml. of ether. The mixture is cooled and 117.1 g. (0.6 mol.) of 2-bromo-2-methylpropionic acid ethyl ester is added. The reaction mixture is warmed for two hours, then stirred at ambient temperature for 48 hours. Acetic acid and water are added, the layers are separated and the ethereal phase is washed with water, dried (MgSO4) and evaporated to dryness to give diethyl 2-(1-carbethoxy-l-methylethyl)malonate.
When an equivalent amount of the following bromo-esters:
3-bromo-2,2-dimethylpropionic acid methyl ester 5-bromo-2-methylvaleric acid methyl ester

2-bromo-2-ethylbutyric acid methyl ester : is used in place of 2-bromo-2-methylpropionic acid e-thyl ester in the reaction with diethylmalonate, the substituted malonates listed below are obtained:
diethyl 2-(2-carbomethoxy-2-methylpropyl)-malonate diethyl 2-(4-carbomethoxypentyl)malonate ~ diethyl 2-(1-carbomethoxy-1-ethylpropyl)malonateO
: Substitution of a substituted malonate named hereinabove in place of diethyl 2-(carbomethoxyethyl)malonate, as a starting material in the p~ocedure of Example 5 followed by the subsequent syn~hetic steps described therein, gives the following triazole thiols as products:
4-(l-carboxy-l-methylethyl)-1,2,3-triazole-5-thiol 4-(2-carboxy-2-methylpropyl)-1,2,3-triazole-5-thiol 4-(4-carboxypentyl)-1,2,3-triazole-5-thiol 4-(1-carboxy-l-ethylpropyl)-1,2,3-triazole-5-thiol.
Use of a substituted triazole thiol sodium salt, prepared as described above, in the procedure of Example l in place of 3-carboxymethyl-1,2,4-triazole-5-thiol sodium salt gives the following 7-(l-tetrazolylacetamido)-3-(sub-stituted triazolyl)thiomethyl cephalosporin compounds:
. 7-(l-tetrazolylacetamido)-3-[4-.. (l-carboxy-l-methylethyl)-1,2,3-triazol-5-ylthiomethyl]-3-cephem-4-carboxyli.c acid ; 7-(1-tetrazolylacetamido)-3-[4-(2-carboxy-2-methylpropyl)-1,2,3-triazol-5-ylthiomethyl]-3-cephem-4-carboxylic acid ~0~s~

7-(1-tetrazolylacetamido)-3-[4-(4-carboxypentyl)-1,2,3-triazol-5-ylthiomethyl]-3 cephem-4-carboxylic acid 7-(1-tetrazolylacetamido)-3-~4-(1-carboxy 1-ethylpropyl)-1,2,3-triazol-5-ylthio-; methyl]-3-cephem-4-carboxylic acid.
; EXAMPLE 8 7-(1-Tetrazolylacetamido)-3-(4-N-methylcarbamoyl-1,2,3-triazol-5-ylthiomethyl)-3-cephem-4-carboxylic acid A solution of 7.2 g. (44 mmol.) of l,l-carbonyl-diimidazole in 110 ml. of dry tetrahydrofuran and 20 ml. of dimethylformamide is added to a solution of 7.7 g. (43.8 mmol.) of 4-carboxy-1,2,3-triazol-5-thiol in 110 ml. of ary tetra-; hydrofuran and 20 ml. of dimethylformamide. Tetrahydrofuran saturated with methylamine is added and the reaction mixture ` is stirred at 25 for 12 hours. The mixture is evaporated ~ to dryness, the residue is diluted with 200 ml. of water and ; the resulting solution is adjusted to pH 2-3 by addition of sulfuric acid. The aqueous solution is lyophilized and the residue extracted with acetone to give 4-N-methylcarbamoyl-1,2,3-triazole-5-thiol.
Reaction of 4-N-methylcarbamoyl-1,2,3-triazole-5-~- thiol sodium salt, prepared as described above, with 7-(1-tetrazolylacetamido)cephalosporanic acid as described in the procedure of Example 1 gives the title compound.

When ethylamine, propylamine or butylamine is sub-stituted for methylamine in the procedure of Example 8, the . ., following triazole thiols are prepared:
....

~ 30 4-N-ethylcarbamoyl-1,2,3-triaæole-5-thiol ,,' 4-N-propylcarbamoyl-1,2,3-triazole-5-thiol 4-N-butylcarbamoyl-1,2,3-txiazole-5-thiol '' .

:
:, . .
.~ .
"' "' ' ' "

1~)7Z~3 Reaction of the sodium salt of a triazole thiol listed above, prepared as described in Example 1, with 7~ tetrazolylacetamido)cephalosporanic acid or 7-(2-thienylacetamido)cephalosporanic acid as described herein-above, gives the following compounds of this invention:
7-(l-tetrazolylacetamido)-3-(4-N-ethylcarbamoyl-1,2,3-triazol-5-ylthiomethyl)-3-cephem-4-carboxylic acid 7-(1-tetrazolylacetamido)-3-(4-N-propylcarbamoyl-1,2,3-triazol-5-ylthiomethyl)-3-cephem-4-carboxylic acid 7-(1-tetrazolylacetamido)-3-(4-N-butylcarbamoyl-1,2,3-triazol-5-ylthiomethyl)-3-cephem-4-carboxylic acid 7-(2-thienylacetamido)-3-(4-N-ethyl-carbamoyl-1,2,3-triazol-5-ylthiomethyl)-3-cephem-4-carboxylic acid 7-(2-thienylacetamido)-3-(4-N-propyl-carbamoyl-1,2,3-triazol-5-ylthiomethyl)-3-; 20 cephem-4-carboxylic acid 7-(2-thienylacetamido)-3-(4 N butyl-carbamoyl-1,2,3-triazol-5-ylthiomethyl)-3-cephem-4-carboxylic acid E~AMPLE lO

7-(1-Tetrazolylacetamido)-3-(5-N,N-dimethylcarbamoylmethyl - 1,2,4-triazol-3-ylthiomethyl)-3-cephem-4-carboxylic acid To a solution of 5.34 g~ (33.8 mmol.) of 3-carboxymethyl-1,2,4~triazole-5-thiol in 75 ml. of dry tetrahydrofurarl and 20 ml. of dry dimethylformamide is slowly added a solution of 5.49 g. (33.9 mmol.) of 1,1-carbonyldiimidazole in 95 ml. of dry dimethylformamide.
-' . , . , . . , ' .

., !
~ 3 The reaction mixture is stirred for 35 minutes, then 250 ml.
of tetrahydrofuran saturated with dimethylamine is added to the suspension and it is stirred at 25 for 12 hours. The mixture is concentrated to about 200 ml. and tetrahydrofuran and ether are added. The precipitate is collected by filtra-tion and dissolved in 170 ml. of water. The aqueous solution is acidified to pH 2.0 by addition of 6N sulfuric acid and extracted with ethyl acetate. The ethyl acetate solution is - evaporated to dryness and triturated with ether to give 3-N, N-dimethylcarbamoylmethyl-1,2,4-triazole-5-thiol.
Reaction of 3-N,N-dimethylcarbamoylmethyl-1,2,4-triazole-5-thiol sodium salt, prepared as described in `~ Example 1, with 7-(1-tetrazolylacetamido)cephalosporanic acid as described therein gives the title compound.
E ~ ~LE 11 When diethylamine, dipropylamine or dibutylamine is substituted for dimethylamine in the procedure of Example 10, the following triazole thiols are prepared:

3-N,N-diethylcarbamoylmethyl-1,2,4-triazole-5-thiol 3 N,N dipropylcarbamoylmethyl-1,2,4-triazole-5-thiol 3-N,N-dibutylcarbamoylmethyl-1,2,4-triazol-5-thiol Reaction of the sodium salt of a triazole thiol listed above, prepared as described in Example 1, with . 7-tl-tetrazolylacetamido)cephalosporanic acid or 7-(2-thienylacetamiao)cephalsoporanic acid as described above ., gives the following compounds of this invention, respectively:

`~^
' ." -- 19 --.. ", ' ' ' .
,' ' ' 1 l)~2Sf~3 7-(1-tetrazolylacetamido)-3-(5-N,N-diethylcarbamoylmethyl-1,2,4-triazol-3-ylthio-methyl)-3-cephem-4-carboxylic acid 7-(1-tetrazolylacetamido)-3-(5-N,N-dipropylcarbamoylmethyl-1,2,4-triazol-3-ylthiomethyl)-3-cephem-4-carboxylic acid 7-(1-tetrazolylacetamido)-3-(5-N,N-dibutylcarbamoylmethyl-1,2,4-triazol-3-ylthio-methyl)-3-cephem-4-carboxylic acid 7-(2-thienylacetamido)-3-(5-N,N-diethylcarbamoylmethyl-1,2,4-triazol-3-ylthio-methyl)-3-cephem-4-carboxylic acid 7-(2-thienylacetamido)-3-(5-N,N-dipropylcarbamoylmethyl-1,2,4-triazol-3-ylthiomethyl)-3-cephem-4-carboxylic acid 7-(2-thienylacetamido)-3-(5-N,N-dibutylcarbamoylmethyl-1,2,4-triazol-3-ylthio-methyl)-3-cephem-4-carboxylic acid.

7-(2 Thienylacetamido?-3-(1-carboxymethyltetrazol-5-ylthio-methyl?-3-cephem-4-carboxylic acid Glycine (60 g., 0.8 mol.) was added to a cooled (5-10) solution of 89.6 g. (1.6 mol.) of po-tassium hydroxide in 200 ml. of water. After complete dissolution 60.8 g.
; (0.8 mol.) of carbon disulfide was added and the reaction mixture was stirred at 25 for three hours. A solution of 113.6 g. (0.8 mol.) of methyl iodide in 200 ml. of ethanol was added while malntaining the temperature at 25-30~. The reaction mixture was stirred for two hours a-t 25, then con-centrated in vacuo and the remaining aqueous phase was made basic (pH 8.0) with aqueous sodium carbona-te and extrac-ted t - 20 -' ~ ~U~7~

with ether. The aqueous phase was acidified to pH 2.5 with dilute hydrochloric acid and extracted with ethyl acetate.
The extract was concentrated in vacuo and the residue re~
crystallized from toluene to give methyl carboxymethyldithio-carbamate.
A mixture of 16.5 g. (0.1 mol.) of methyl carboxy-methyldithiocarbamate and 14.3 g. (0.22 mol.) of sodium azide in 150 ml. of water was heated at 56 for 12 hours. The ; reaction mixture was extracted with ether, then acidified to pH 1.5 with dilute hydrochloric acid and ex-tracted with e-thyl acetate. The extract was dried (MgSO4) and evaporated to dryness to give l-carboxymethyltetrazole-5-thiol, m.p. 178-179.
l-Carboxymethyltetrazole-5-thiol was also prepared by refluxing a mixture of 45.95 g. (0.316 mol.) of ethyl isothiocyanoacetate and 30.8 g. (0.475 mol.) of sodium azide in 500 ml. of water for 2.75 hours. Ethyl acetate (400 ml.) was added to the cooled reaction mixture and it was acidified to pH 1.9 with 3N hydrochloric acid. The layers were separated, ~- 20 the aqueous phase was extracted three times with ethyl acetate and the combined extracts were dried (MgSO4) and evaporated to dryness to give a residue which was chromatographed on silica gel with 17:3:2 chloro-form-isopropanol-formic acid to ` give the tetrazole thiol.
A mixture of 2.94 g. (10 mmol.) of 7-aminocephalo-; sporanic acid sodium salt, 2.40 g. (15 mmol.) of 1-carboxy-methyltetrazole-5-thiol and 2.52 g. (30 mmol.) of sodium bicarbonate in 40 ml. of water was heated at 70 for four hours. The reaction mixture was cooled (ice bath), acidified to pH 1.8 with 3N hydrochl~ric acid and evaporated to dryness in vacuo to give 7-amino-3-(1-carboxymethytetrazol-5-ylthio-,.
~ 21 -.:

~ 3 methyl)-3-cephem-4-carboxylic acid.
To a solution of 1.86 g. (5 mmol.) of 7-amino-3-(l-carboxymethyltetrazol-S-ylthiomethyl)-3-cephem-4-carboxylic acid and 1.68 g. (0.02 mol.l of sodium bicarbonate in 60 ml.
of water and 50 ml. of acetone at -15 was added a solution of 0.80 g. (5 mmol.) of 2-thiophene acetyl chlorlde in 50 ml.
of acetone. The reaction mixture was stirred and the tempera-ture maintained below -10 during addition. The mixture was stirred for 3.5 hours, then the acetone was removed ln vacuo and the~aqueous residue was extracted with ethyl acetate.
Ethyl acetate was added to the aqueous phase and it was acidified to pH 1.5 by addition of 3N hydrochloric acid. The layers were separated and the aqueous phase was extracted with ethyl acetate. The extract was dried (MgSO4) and evaporated i to dryness to give a residue, which was chromatographed on silica with 8:2:1 chloroform-isopropanol-acetic acid as - eluent. The product was dissolved in ethyl acetate and triturated with hexane to give the title compound.
C17H16N6O6S3 0.25 C4 8 2 Calculated: 41.69% C; 3.49% H; 16.20% N
Found: 41.51% C; 3.54% H; 15.74% N
~ The title compound is dissolved in methanol and the ; methanol solution is treated with 0.196N sodium methoxide in methanol. The methanol is removed in vacuo and the residue .- .
is dissolved in a minimum amount of water to which isopropanol ; is added to give 7-(2-thienylacetamido)-3-(1-carboxymethyl-tetrazol-5-ylthiomethyl)-3-cephem-4-carboxylic acid sodium salt.

30 7-(2-Thienylacetamido)-3-(1-N-methylcarbamoylmethyltetrazol-.
5-ylthiomethyl)-3 cephem-4-carboxylic acid . ~
A solution of 7.2 g. (44 mmol.) of l,l-carbonyl-~'7~

diimidazole in 110 ml. of dry tetrahydrofuran and 20 ml. of dimethylformamide was added to a solution of 7.0 g. (43.8 mmol.) of l-carboxymethyltetrazole-5-thiol in 110 ml. of dry tetrahydro-furan and 20 ml. of dimethylformamide. Tetrahydrofuran saturated with methylamine was added and the reaction mixture was stirred at 25 for 12 hours. The mixture was evaporated to dryness ln vacuo, the residue was diluted with 200 ml. of water and the resulting solution was adjusted to pH 2-3 by addition of 3N
hydrochloric acid. The aqueous solution was extracted with ethyl acetate and the extract was dried (MgSO4) and evaporated to dry-ness. Ethyl acetate (15 ml.) and ether (10 ml.) were added to the residue and it was cooled to induce crystallization of l-N-methylcarbamoylmethyltetrazole-5-thiol, m.p. 137-140.
Reaction of l-N-methylcarbamoylmethyltetrazole-5-thiol, 7-aminocephalosporanic acid sodium salt and sodium bicarbonate as described in the procedure of Example 12 gives 7-amino-3-(1-N-~` methylcarbamoylmethyl-tetrazol-5-ylthiomethyl)-3-cephem-4-carboxy-lic acid.
Substitution of 7-amino-3-(1-N-methylcarbamoylmethyl-tetrazol-5-ylthiomethyl)-3-cephem-4-carboxylic acid in -the pro-cedure of Example 12 for 7-amino-3-(1-carboxymethyltetrazol-5-ylthiomethyl)-3-cephem-4-carboxylic acid while maintaining the pH at 7~6 - 7.8 throughout the reaction by addi-tion of solid sodium bicarbonate gives the title compound.
: EX~MPLE 14 7-(2-Thienylacetamido)-3-(1-N,N-dimethylcarbamoylmethyltetrazol-5-ylthiomethyl)-3-cephem-4-carboxylic acid To a solution of 5.42 g. (33.8 mmol.) of l-carboxymethyl-tetrazole-5-thiol in 75 ml. of dry tetrahydrofuran and 20 ml. of dry dimethylformamide was s:Lowly added a solution of 5.49 g.
(33.9 mmol.) of l,l-carbonyldiimidazole in 95 ml. of dry dimethyl-formamide. The reaction mixture was stirred for 35 minutes, then ~ 23 -250 ml. of tetrahydrofuran saturated with dimethylamine was added to the suspension and it was stirred at 25 for 12 hours. The mixture was concentrated to about - 23a -:~a3';'2~43 200 ml. and tetrahydrofuran and ether were added. The pre-cipitate was collected by filtration and dissolved in 170 ml.
of water. The aqueous solution was acidified to pH 2.0 by addition of 6N sulfuric acid and extracted with ethyl acetate.
The ethyl acetate solution was evaporated to dryness and the residue was triturated with ether to give l-N,N-dimethyl-carbamoylmethyltetrazole-5-thiol, m.p. 190-200 (dec.).
Reaction of l-N,N-dimethylcarbamoylmethyltetrazole-5-thiol with 7-aminocephalosporanic acid sodium salt as described in Example 13 followed by reaction of the 7-amino-3-(1-N,N-dimethylcarbamoylmethyltetrazol-5-ylthiomethyl)-3-cephem-4-carboxylic acid thus formed with 2--thiophene acetyl chloride as described in Example 12 gives the title compound.

~,- 7-(2-Thienylacetamido3-3-(1-carbamoylmethyltetrazol-5-ylthiomethyl)-3-cephem-4-carboxylic acid l-Carboxymethyltetrazole-5-thiol and l,l-carbonyl-diimidazole were reacted as described in Example 13. To the reaction mixture was added tetrahydrofuran saturated with ; 20 dry ammonia gas. The resulting suspension was stirred for 2.5 hours and the solid which formed was collec-ted by filtration, washed with tetrahydrofuran and dissolved in methanol.
Amberlite IR-120 ion-exchange resin (50 g.) was added and the suspension was stirred for 15 minutes. The resin was then - removed and washed with absolute methanol. The methanol solu-tion was evaporated to dryness to give l-carbamoylmethyltetra-zole-5-thiol, m.p. 200 (dec.).
l-Carbamoylmethyltetrazole-5-thiol is reacted with 7-aminocephalosporanic acid sodium salt to give 7-amino-3-(1-carbamoylmethyltetrazol-5-ylthiomethyl)-3-cephem~4-carboxy-. . .
lic acid, which upon reaction with 2 thiophene acetyl chloride as described in Example 12, gives -the -ti-tle compound.

' .', ~

~'7~5'~

~EXAMPLE 16 7-(2-Thienylacetamido)-3-[1-(2-carboxyethyl)tetrazol-5-ylthiomethy-1]-3-cephem-4-carboXy-lic acid 3-Alanine (17.8 g., 0.2 mol.) was added to a solu-tion of 22.4 g. (0.4 mol.) of potassium hydroxide in 500 ml.
of water at 25. Carbon disulfide (12.2 ml., 0.2 mol.) was added and the reaction mixture was refluxed for three hours.
The mixture was cooled, 28.4 g. (0.2 mol.) of methyl iodide and 500 ml. of ethanol were added and the resulting mixture was stirred for 30 minutes. The precipitate was collected by filtration, the filtrate was cancen-trated and the aqueous residue was combined with the solid material and brought to pH 8.5-9 by addition of 10% aqueous sodium hydroxide. The resulting suspension was extracted with ethyl acetate and the extract discarded. Ethyl acetate was added to the aqueous phase which was then acidified to pH 1.5 with 6N hydrochloric acid. The layers were separated and the aqueous phase was extracted with ethyl acetate. The ethyl acetate extracts were combined, dried (MgSO4) and evaporated to dryness to give methyl 2-carboxyethyldithiocarbamate.
To a mixture of 25.37 g. (0.143 mol.) of methyl 2-carboxyethyldithiocarbamate and 5.6 g. (0.143 mol.) of sodium hydroxide in 210 ml. of water was added 9.25 g.
- (0.143 mol.) of sodium azide. The reaction mixture was re-- fluxed for one hour then cooled, 100 ml. of ether was added ; and the mix-ture was acidified to pH 1.7O The layers were separated, the aqueous phase was extracted with ether and the combined extracts were dried (MgSO~) and evaporated to dryness to give a residue which was recrystallized from acetone-chloroform to give 1-(2-carboxyethyl)tetrazole-5-thiol, m.p. 158-160.

' To a solution of 4.18 g. (0.01 mol.) of 7-(2- ~
thienylacetamido)cephalosporanic acid sodium salt in 50 ml.
of water was added 2.51 g. (0.015 mol.) of 1-(2-carboxyethyl)-tetrazole-5-thiol and 1.26 g. (0.015 mol.) of sodium bicar-bonate. Additional amounts of sodium bicarbonate were added to bring the pH of the reaction mixture to 7.0 and the mixture was heated at 69 for five hours. The reaction mixture was cooled, acidified to pH 6.0 by addition of dilute hydrochloric acid and extracted with ethyl acetate. The aqueous phase was acidified to pH 2.0, extracted with ethyl acetate and the extract was evaporated to dryness to give a residue which was chromatographed on silica with 90:10:3 chloroform-methanol-acetic acid as eluant to give the title compound.
The title compound was dissolved in ethyl acetate and triethylamine was added to form the corresponding tri-ethylamine salt.

Calculated: 46.43% C; 5.95% H; 15.79% N
Found: 46.51% C; 5.83% H; 14.98% N

7-(2-Thienylacetamido)-3-[1-(2-carbamoylethyl)tetrazol-5-ylthiomethyl]-3-cephem-4-carboxylic acid When an equivalent amount of 1-(2-carboxyethyl)-tetrazole-5-thiol was substituted in the procedure of Example 15 for 1-carboxymethyltetrazole-5-thiol, 1-(2 carbamoylethyl)-; tetrazole-5-thiol was ob-tained, m.p. 181--]82 (dec.).
~ solution of 1-(2-carbamoylethyl)tetrazole-5-thiol (10.4 g., 0.06 mol.) in 120 ml. of acetone was added to a warm (45) solution of 10.9 g. (0.04 mol.) of 7-aminocephalo-sporanic acid in a mixture of 220 ml. of water, 50 ml. oE
acetone and 8.4 g. (0.01 mol.) of sodium bicarbonate. The :' , -~6 temperature was raised to 65 and the pH maintained at 7.4-~7.6 by addition of aqueous sodium carbonate solution. After three hours, the reaction mixture was cooled to 10 and adjusted to pH 3.5 by addition of dilute hydrochloric acid. The re-sulting solid was collected by filtration, washed with water and acetone and suspended in 95 ml. of 1.5N hydrochloric acid.
The acid suspension was stirred at 25 for five hours, filtered and the pH of the filtrate was adjusted to 3.5 by addition of solid sodium bicarbonate. The solid was collected by filtra-tion and washed with water and acetone to give 7-amino-3-[1-(2 carbamoylethyl)tetrazol-5-ylthiomethyl-]-3-cephem-4-carboxylic acid.
7-Amino-3-[l-(2-carbamoylethyl)tetrazol-5-ylthio-methyl]-3-cephem-4-carboxylic acid and 2-thiophene acetyl chloride were reacted according to the procedure of Example 13 to give the title compound.
The title compound was dissolved in methanol and sodium methoxide solution was added until pH 6.9. Evaporation of the methanol gave the title compound as its sodium salt.
18 18N7O5S3 Na 0.5 H2O
Calculated: 39.99% C; 3.54% H; 18.13% N
Found: 39.92% C; 3.48% H; 17.60% N
7-(2-Thienyl)-3-[l-(2-carbamoylethyl)tetrazol-5-yl-thiomethyl]-3-cephem~4-carboxylic acid sodium salt is converted to the title compound by methods previously described.

7-(1-Tetrazolylacetamido)-3-(1-carboxymethylte-trazol-5-ylthiomethyl)-3 cephem-4-carboxylic aci_ 7-(1-Tetrazolylacetamido)cephalosporanic acid (1.41 g., 3.7 mmol.) and 0.96 g. (6.0 ~nol.) of l-carboxymethyltetra-zole-5-thiol were added to a solution of 0.815 g. (9.7 mmol.) . . .
.

, .

%~

of sodium bicarbonate in 25 ml. of water. An additional 0.5`04 g. (6.0 mmol.~ of sodium bicarbonate was added to bring the pH
to 6.7 and the reaction mixture was heated at 63 for five hours while maintaining the pH at 6.6-6.8 by addition of acetic acid. The mixture was cooled, acidified to pH 1.9 by addition of 3N hydrochloric acid and extracted with ethyl acetate. The extract was dried (Na2SO4) and evaporated to dryness. The residue was dissolved in ethyl acetate and ether and hexane - were added to precipitate the title compound.
C14EIl~Nl0o6s2 0.6 C4 8 2 Calculated: 36.79% C; 3.53% H; 26.16% N
Found: 36.28% C; 3.50% H; 25.85% N
EXAr~PLE 19 7-(1-Tetrazolylacetamido)-3-[1 (2-carboxyethyl)tetrazol-_ylthiomethyl]-3-cephem-4-carboxylic acid When an equivalent amount of 1-(2-carboxyethyl)-tetrazole-5-thiol was substituted in the procedure of Example ` 18 in place of 1-carboxymethyltetrazole-5-thiol, the title compound was obtained.

Calculated: 36.28% C; 3.24% H; 28.21% N
Found: 35.80% C; 3.67% H; 24.16% N
EXA~PLE_20 7-(1-Tetrazolylacetamido)-3-[1-(3-carboxypropyl)tetrazol 5-ylthiomethyl]-3-cephem-4-carboxyllc acid When an equivalent amount of 4-aminobutyric acid was substituted in the procedure of Example 16 Eor ~-alanine, ~ methyl 3-carboxypropyldithiocarbamate was prepared.
- Reaction of methyl 3-carboxypropyldithiocarbamate with sodium azide and sodium hydroxide as described in Example ; - 28 -' 16 gave 1-(3-carboxypropyl)tetrazole-5-thiol, m.p. 99-101.
Substitution of an equivalent amount of 1-(3-carboxypropyl)te-trazole-5-thiol in the procedure of Example 18 in place of 1-carboxymethyltetrazole-5-thiol gave the title compound.
6Nloo6s2 2 Na 2H2O 0.5 C3H8O
Calculated: 33.87% C; 3.89% H; 22.57% N
Found: 34.39% C; 3.46% H; 22.20% N

, 10 7-(2-Thienylacetamido-3-[1-(3-carbamoylpropyl)tetrazol-5-ylthiomethyl]-3-cephem-4-carboxylic acid When an equivalent amount of 1-(3-carboxypropyl)-tetrazole-5-thiol was substituted in the procedure of Example 15 for 1-carboxymethyltetrazole-5-thiol, 1-(3-carbamoylpropyl)-tetrazole-5-thiol was obtained, m.p. 133-136.
Reaction of 1-(3-carbamoylpropyl)tetrazole-5-thiol and 7-aminocephalosporanic acid as described in the procedure of Example 17 gives 7-amino-3-[1-(3~carbamoylpropyl)tetrazol-5-ylthiomethyl]-3-cephem-4-carboxylic acid.
/-Amino-3-[1-(3-carbamoylpropyl)tetrazol-5-ylthio-methyl]-3-cephem-4-carboxylic acid and 2-thiophene acetyl chloride are reacted according to the procedure of Example 13 to give the title compound.

7-(1-Tetrazolylacetamido)-3-[1-(5-carboxypentyl)tetrazol-5-ylthiomethy]-3-cephem-4-carboxylic acid Substitution of an equivalent amount of 6-amino-caproic acid in the procedure of Example 16 for ~-alanine gave methyl 5-carboxypentyldithiocarbamate.
~ 30 Reaction of methyl 5-carboxypentyldi-thiocarbamate - with sodium azide and sodium hydroxide as described in Example s~:~

16 gave 1-(5-carboxypentyl)tetrazole-5-thiol, m.p. 100-100.5.
Substitution of an equivalent amount of 1-(5- -carboxypentyl)tetrazole-5-thiol in the procedure of Example 18 in place of 1-carboxymethyltetrazole-5-thiol gave the title compound.
C18H22N10O6S2 0.33 CH40 Calculated: 40.09% C; 4.28% H; 25.50% N
Found: 40.29% C; 4.18% H; 25.78% N

7-(1-Tetrazolylacetamido)-3-rl-(5-carbamoylpentyl)tetrazol 5-ylthiomethyl]-3-cephem-4-carboxylic acid 1-(5-Carboxypentyl)tetrazole-5-thiol (6.0 g., 28 mmol.) was slowly dissolved in 20 ml. of thionyl chloride and the mixture was stirred at 25 for 1.5 hours. Evaporation of the reaction mixture to dryness gave a residue which was dis-; solved in 30 ml. of tetrahydrofuran. The tetrahydrofuran solu-tion was added to a cold mixture of 60 ml. of ammonium hydroxide and 30 ml. of tetrahydrofuran and the resulting mixture was stirred at 25 for two days. The mixture was extracted with ethyl acetate. The aqueous phase was acidified to pH 1 by addition of 6N hydrochloric acid to precipitate 1-(5-carbamoyl-pentyl)tetrazole-5-thiol, m.p. 155-157.
Reaction of 7-(1-tetrazolylacetamido)cephalosporanic acid, 1-(5-carbamoylpentyl)tetrazole-5-thiol sodium salt, pre-pared from 1-(5-carbamoylpentyl)tetrazole-5-thiol as described in Example 1, and sodium bicarbonate according to the procedure - of Example 1, gives the title compound.
EXA~PLE 24 7-(2-Thienylacetamido)-3-[1-(10 carbamoyldecyl)tetrazol-5-ylthiomethyl]-3-cephem-4-carboxylic acid , l-(10-Carbamoyldecyl)tetrazole-5-thiol was prepared ' , 25'~

from 1-(10-carboxydecyl)tetrazole-5-thiol by the procedure described in Example 15, m.p. 112-114.
To a solution of 0.084 g. (1.0 mmol.) of sodium bicarbonate in 11 ml. of water was added 0.285 g. (1.0 mmol.) of l-(10-carhamoyldecyl)tetrazole-5-thiol. The reaction mixture was heated to ca. 66, 0.250 g. (0.6 mmol.) of 7-(2-thienyl-acetamido)cephalosporanic acid sodium salt was added and the heating was continued for 5.5 hours. The reaction mixture was chromatographed on XAD-4 resin with water as the eluant. The product-containing fractions were evaporated to dryness and lyophilized and the residue was chromatographed on silica with 9:1:0.5 chloroform-isopropanol-formic acid as eluant to give the title compound.
The title compound was converted to the correspond-ing sodium salt as previously described.

Calculated: 47.79% C; 5.36% H; 15.01% N
Found: 48.17% C; 5.37% H; 14.52% N

7-(1-Tetrazolylacetamido)-3-[1-(10-carboxydecyl)tetrazol-5-ylthiomethyl]-3-cephem-4-carboxylic ac d A suspension of 56 g. (1.0 mol.) of potassium hydroxide and 100 g. (0.5 mol.) of ll-aminoundecanoic acid in 170 ml. of water was stirred for 30 minutes at 25 then 40 g. (0.52 mol.) of carbon disulfide and 80 ml. of ethanol were added and the reaction mixture was stirred at 25 for 12 hours. The mixture was refluxed gently for two hours and ; cooled. Methyl iodide (71 g., 0.3 mol.) and 130 ml. of ethanol were added to the mixture and it was stirred at 25 , 30 for 12 hours. The mixture was evaporated to remove the ethanol and the solid residue was collected by filtration to yive ; me-thyl 10-carboxydecyldithiocarbamate potassium salt.

.
.

!~ .
; Methyl 10-carboxydecyldithiocarbamate (28 g., 0.096 mol.), obtained from the potassium salt as described above, was reacted with 6.5 g. ~0.1 mol.) of sodium azide according to the procedure described in Example 16. Acidification upon work-up gave l-(10-carboxydecyl)tetrazole-5-thiol as a white precipitate, m.p. 95-98.
l-(10-Carboxydecyl)tetrazole-5-thiol, 7-(1-tetrazolyl-acetamido)cephalosporanic acid sodium salt and sodium bicarbonate were reacted as described in Example 24 to give the title com-pound.

7-(1-Tetrazolylacetamido)-3-[1-(2-carboxy-1-methylethyl)-tetrazol-5-ylthiomethyl]-3-cephem-4-carboxylic acid ... .. ... _ .
Substitution of an equivalent amount of 3-aminobutyric acid in the procedure of Example 16 for ~-alanine gave methyl (2-carboxy-1-methyl)ethyldithiocarbamate.
Treatment of methyl (2~carboxy-1-methyl)ethyldithio-carbamate with sodium azide also as described in Example 16 gave 1-(2-carboxy-1-methylethyl)tetrazole-5-thiol, m.p. 169-172.
7-(1-Tetrazolylacetamido)cephalosporanic acid and 1-(2-carboxy-1-methylethyl)tetrazole-5-thiol are reacted in the presence of excess sodium bicarbonate as described in Example 16 to give the title compound.

When an equivalent amount of an amino acid listed be-low:
alanine 2-aminobutyric acid 2-aminovaleric acid 2-aminohexanoic acid , ~ 3 is used in the procedure of Example 16 in place of ~-alanine and the resulting dithiocarbamates are treated with sodium azide as described therein, the following substituted tetrazole thiols are obtained:
l-(l-carboxyethyl)tetrazole-5-thiol l-(l-carboxypropyl)tetrazole-5-thiol l-(l-carboxybutyl)tetrazole-5-thiol l-(l-carboxypentyl)tetrazole-5-thiol Reaction of a tetrazole thiol listed above wlth 7-(1-tetrazolylacetamido)cephalosporanic acid as described hereinabove gives the following compounds of this invention:
7-(1-tetrazolylacetamido)-3-[1-(1-carboxyethyl)-tetrazol-5-ylthiomethyl]-3-cephem-4-carboxylic acid 7-(1-tetrazolylacetamido)-3-[1-(1-carboxypropyl)-tetrazol-5-ylthiomethyl]-3-cephem-4-carboxylic acid 7-(1-tetrazolylacetamido)-3-[1-(1-carboxybutyl)-tetrazol-5-ylthiomethyl]-3-cephem-4-carboxylic acid 7-(1-tetrazolylacetamido)-3-[1-(1-carboxypentyl)-tetrazol-5-ylthiomethyl]-3-cephem-4-carboxylic acid.
Likewise, reaction of a substituted tetrazole thiol listed a~ove with 7-(2-thienylacetamido)cephalosporanic acid according to theprocedures described herein gives the corres-ponding 7--(2--thienylacetamido)-3-(1-carboxyalkylte-trazol-5-ylthiomethyl)-3-cephem-4-carboxylic acids.

The following compounds were prepared according -to procedures described hereinabove:
7--(2-thienylacetamido)-3-[1-(3-carboxypropyl)tetrazol-i 5-ylthiomethyl]~3-cephem-4-carboxylic acid lgH18N6S3O6 2 Na 2 H2O

Calculated: 37.75% C; 3.67% H; 13.89% N
Found: 38.25% C; 3.61% H; 13.49% N

^--~
:
~ 5'-~
7~(2-thienylacetamido-3-[1-(5-carboxypentyl)-tetrazol-5-ylthiomethyl]-3-cephem-4-carboxylic acid 21H22N66S3 2 Na 0.75 H2O
Calculated: 41.34% C; 3.88% H; 13.77% N
Found: 41.30% C; 3.76% H; 13.58% N

An injectable pharmaceutical composition is formed by adding sterile water or sterile saline solution (2 ml.) to 500 mg. of 7-(2-thienylacetamido)-3-~1-(2-carbamoylethyl)-; 10 tetrazol-5-ylthiomethyl]-3-cephem-4-carboxylic acid, sodium salt.
Pharmaceutical compositions of the other antibacterial compounds disclosed above may be form~lated in a similar manner.

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Claims

The embodiments of the invention in which an exclusive property or privilege is claimed are defined as follows:

1. A process for preparing a compound of the formula:

in which:
R1 is thienyl or tetrazolyl; and Het is selected from the group consisting of:

' and in which each individual R2 is hydrogen or lower alkyl, n is zero to ten, m is one to ten and R3 is hydroxy, amino, lower alkylamino or di(lower)alkylamino, or a non-toxic pharmaceutically acceptable salt thereof, comprising acylating a compound of the formula:

where Het is defined as above and R4 is hydrogen or a protecting ester group, with thienyl or tetrazolyl acetic acid or an activated derivative thereof, or reacting a compound of the formula:

where R1 is defined as above or a salt thereof, with a compound of the formula:

, or where R2, R3, n and m are defined as above or a salt thereof, followed by removal of the protecting group when necessary, acidifying when necessary and optionally converting the product to a non-toxic pharmaceuti-cally acceptable salt.

2. A process as claimed in claim 1 in which Het is triazolyl.

3. A process as claimed in claim 1 in which Het is tetrazolyl.

4. A process as claimed in claim 2 in which n is zero to five, R2 is hydrogen and R3 is hydroxy or amino.

5. A process as claimed in claim 3 in which m is one to five, R2 is hydrogen and R3 is hydroxy or amino.

6. A process as claimed in claim 3 for preparing 7-(2-thienylacetamido)-3-[1-(10-carbamoyldecyl)tetrazol-5-ylthiomethyl]-3-cephem-4-carboxylic acid comprising reacting 7-(2-thienylacetamido)cephalosporanic acid sodium salt with 1-(10-carbamoyldecyl)tetrazole-5-thiol.

7. A process as claimed in claim 4 for preparing 7-(1-tetrazolylacetamido)-3-(5-carboxymethyl-1,2,4-triazol-3-ylthiomethyl)-3-cephem-4-carboxylic acid comprising reacting 7-(1-tetrazolylacetamido)cephalo-sporanic acid with 3-carboxymethyl-1,2,4-triazole-5-thiol sodium salt and then acidifying.

8. A process as claimed in claim 5 for pre-paring 7-(2-thienylacetamido)-3-(1-carboxymethyltetrazol-5-ylthiomethyl)-3-cephem-4-carboxylic acid comprising acylating 7-amino-3-(1-carboxymethyltetrazol-5-ylthio-methyl)-3-cephem-4-carboxylic acid with 2-thiophene acetyl chloride.

9. A process as claimed in claim 5 for pre-paring 7-(2-thienylacetamido)-3-[1-(2-carbamoylethyl)tet-razol-5-ylthiomethyl]-3-cephem-4-carboxylic acid comprising acylating 7-amino-3-[1-(2-carbamoylethyl)tetrazol-5-ylthiomethyl]-3-cephem-4-carboxylic acid with 2-thio-phene acetyl chloride.

10. A process as claimed in claim 5 for pre-paring 7-(1-tetrazolylacetamido)-3-(1-carboxymethyl-tetrazol-5-ylthiomethyl)-3-cephem-4-carboxylic acid comprising reacting 7-(1-tetrazolylacetamido)cephalo-sporanic acid with 1-carboxymethyltetrazole-5-thiol.

11. A process as claimed in claim 5 for pre-paring 7-(2-thienylacetamido)-3-[1-(2-carboxyethyl)-tetrazol-5-ylthiomethyl]-3-cephem-4-carboxylic acid comprising reacting 7-(2-thienylacetamido)cephalosporanic acid sodium salt with 1-(2-carboxyethyl)tetrazole-5-thiol then acidifying.

12. A process as claimed in claim 5 for preparing 7-(1-tetrazolylacetamido)-3-[1-(2-carboxy-ethyl)tetrazol-5-ylthiomethyl]-3-cephem-4-carboxylic acid comprising reacting 7-(1-tetrazolylacetamido)-cephalosporanic acid with 1-(2-carboxyethyl)tetrazole-5-thiol.

13. A compound of the formula:

in which:
R1 is thienyl or tetrazolyl; and Het is selected from the group consisting of:

and in which each individual R2 is hydrogen or lower alkyl, n is zero to ten, m is one to ten and R3 is hydroxy, amino, lower alkylamino or di(lower)alkyl-amino, or a non-toxic pharmaceutically acceptable salt thereof, when prepared by the process of claim 1 or its obvious chemical equivalent.

14. A compound of claim 13 in which Het is triazolyl, when prepared by the process of claim 2 or its obvious chemical equivalent.

15. A compound of claim 13 in which Het is tetrazolyl, when prepared by the process of claim 3 or its obvious chemical equivalent.

16. A compound of Claim 13 in which Het is triazolyl, n is zero to five, R is hydrogen and R3 is hydroxy or amino, when prepared by the process of claim 4 or its obvious chemical equivalent.

17. A compound of claim 13 in which Het is tetrazolyl, m is one to five, R is hydrogen and R3 is hydroxy or amino, when prepared by the process of claim 5 or its obvious chemical equivalent.

18. The compound 7-(2-thienylacetamido)-3-[1-(10-carbamoyldecyl)tetrazol-5-ylthiomethyl]-3-cephem-4-carboxy-lic acid, when prepared by the process of claim 6 or its obvious chemical equivalent.

19. The compound 7-(1-tetrazolylacetamido)-3-(5-carboxymethyl-1,2,4-triazol-3-ylthiomethyl)-3-cephem-4-carboxylic acid, when prepared by the process of claim 7 or its obvious chemical equivalent.

20. The compound 7-(2-thienylacetamido)-3-(1-carboxymethyltetrazol-5-ylthiomethyl)-3-cephem-4-car-boxylic acid, when prepared by the process of claim 8 or its obvious chemical equivalent.

21. The compound 7-(2-thienylacetamido)-3-[1-(2-carbamoylethyl)tetrazol-5-ylthiomethyl]-3-cephem-4-car-boxylic acid, when prepared by the process of claim 9 or its obvious chemical equivalent.

22. The compound 7-(1-tetrazolylacetamido)-3-(1-carboxymethyltetrazol-5-ylthiomethyl)-3-cephem-4-carboxylic acid, when prepared by the process of claim 10 or its obvious chemical equivalent.

23. The compound 7-(2-thienylacetamido)-3-[1-(2-carboxyethyl)tetrazol-5-ylthiomethyl]-3-cephem-4-carboxylic acid, when prepared by the process of claim 11 or its obvious chemical equivalent.

24. The compound 7-(1-tetrazolylacetamido)-3-[1-(2-carboxyethyl)tetrazol-5-ylthiomethyl]-3-cephem-4-carboxylic acid, when prepared by the process of claim 12 or its obvious chemical equivalent.