Note: Descriptions are shown in the official language in which they were submitted.
., . Los;~71~
ANTIBACTERIAL AGENTS
. The cephalosporins o~ the present in~ént~ion.po~sess the usual attri~utes of such compounds and are particularly :
useful in the txeatment of bacterial in~ections by parenteral
administration.
The cephalosporins are a well-known group of semisyn-
thetic antibacterial agents made originally, for example, by
acylation of the 7-amino group of the nucleus 7-
aminocephalosporanic acid (7-ACA) and later by similar
acylation o~ nuclei derived therefrom, as by modi~ication ~.`
o~ its substituent as the 3-position. Various reviews
have appeared in the scientific literature (e.g. ~-
Cephalosporins and Penicillins - Chemistry and Biology,
edited by Edwin H. Flynn, Academic Press, New York, 1972,
and particularly pages 554-569) and in the patent lit-
erature, e~g. as in U.~. patents 3,6B7,948, 3,741,965,
3,759,904, 3,759,905, 3,766, 175 ~nd 3,776,906 (all
U.S. Class 260-243C). :
Issued patents on 3-thiolated cephalosporins
in which the 7-substituent is ~ -
a) a-Amino-a-phenylacetamido include U.S. . :
3,b41,021, U.S. 3,734,907, U.S. 3,687,948, U.S. ~;
3,741,965, U.S. 3,757,015, U.S. 3,743,644, Japan :
71/24400 (Farmdoc 46374S), Belgium 776,222 (Far~doc
38983Tt U.K. 1,328,340 which includes vaxious substituents
on the benzene ri~g~)', Belgium 772,592 (Farmdoc :
196Y6T; U.S. 3,687,948, 3,734,907 and 3,757,012), :.
West Germany 2,202,274 tFarmdoc 50428T)
., ~ . . .
corresponding to U.S. 3,759,904, Netherlands 7205644 ~
(Farmdoc 76309T; U.S. 3~757n;014); and ~ :
~5~77~ ;:
b~ o-, m- or p~am~noethox~phen~lacetamido as
Net~erlands 72~13968 (Farmdoc 24740tl) corresponding to
U.S. 3,759,905,and
c) o-aminomethylphenylacetamido as Netherlands
72/06326 (Farmdoc 76374T) (which also reviews the older
patent literature concerning substituted 7-phenyl-
acetamidocephalosporanic acids) corresponding bo U.S.
3,766,176 and 3,766,175; and
d) N-(phenylacetimidoyl)aminoacetamido as
U.S. 3,692,779; and
e) ~-amino~ 1,4-cyclohexadienyl)acetamido ~ -
as in Belgium 776,222 ~Farmdoc 38983T; U.R. 1,328,340).
Additional similar disclosures are found in
U.S. 3,692,779 (Belgium 771, 189; Farmdoc 12819T),
Japan 72~05550 (Farmdoc 12921T), Japan 72~05551
(Farmdoc 12922T), U.S. 3,719,673 (Belgium 759,570
Farmdoc 39819S), Belgium 793,311 (Farmdoc 39702U)
and Belgium 793rl91 (Farmdoc 39684U).
Issued disclosures of 3-thiolated cephalosporins
in which the 7-substituent is 7~mandelamido (7-a-
hydroxyphenylacetamido) are found, for example, in
U.S. 3,641,0~1, France 73.10112, U.S, 3,796,801, `
Great Britain 1,328,340 (Farmdoc 38983T), U.S. `!~,;
3,701,775, Japan 4844293 (Farmdoc 55334U) !~and in
Hoover et al., J. Med. Chem. 17(1), 34-41 (1974)
"
and Wick et al., Antimicrobial Ag. Chemo., 1(3),
221-234 (1972).
- 2 -
' ' , . ,
i~S~7~
This invention comprised the acids having the
D configuration in the 7-s~dechain and the formula
-
~1 ~ S~
~- R-fH-C-NH-CH - CH CH2
OH ~C N\ ~C-CH2-S-X
COOH Formula (I) ~ :
wherein X is
~ N=N
N--N
- ~ ~ ~ or ~ - N , and
R is ~.
~ ~ or
and Y is hydrogen, chlorine, bromine, ~luorine, tri- -
fluoromethyl, amino, nitro, hydroxy, loweralkyl of ~ -
1-4 carbon atoms or lower alkoxy of 1-4 carbon atoms and ~
the pharmaceutically acceptable salts of those acids and .
the easily hydrolyzed esters of those acids including
especially the pivaloyloxymethyl, acetoxymethyl, ~ :
acetonyl, phenacyl and methoxymethyl esters and
the silyl esters such as the trimethylsilyl ester.
In the preferred embodiments(~of this invention
R is 2-thienyl, 3-thienyl, phenyl, chlorophenyl, ;'~
bromophenyl, txifluoromethylphenyl, tolyl or
methoxyphenyl. -.
,
.,
':
~)S~7~ :
Such salts include the nontoxic carboxylic acid
salts thereof, including nontoxic metallic sal~s such
as sodium, potassium, calcium and a:Luminum, the
ammonium salt and substituted ammonium salts, e.g.
salts of such nontoxic amines as trialkylamines
including triethylamine, procaine~ dibenzylamine,
N benzyl-beta-phenethylamine, l-ephenamine, N,N'-
dibenzylethylenediamine, dehydroabietylamine, N,N'-
bis-dehydroabietyle~hylenediamine, N-(lower)-alkyl-
piperidine, e.g. N-ethylpiperidine, and other amines
which have been used to form salts with benzyl-
penicillin.
Particularly p~eferred embodiments of this :
inven~ion comprise the acid having the D configuration
in the 7-sidechain and the formulae
~H-C-NH-C~ C~l/ \f N--N ~:
H ~C ~ ~ C-CH2-S ~ OH
1 ~
COOH ;; .
Formula (II) :::
--Cl~-c--NN- ~ C~2 r~
o~C N\ ~1 CH2
100H
Formula (III)
~, ,;' ' ' ' ~ ;
.
~ Z77q:~
and their pharmaceutically acceptable salts and easily
hydrolyzed esters.
Also included in this invention are the compounds
(used as either intermediates or metabolic precursors)
in which the a-hydroxy group is "blocked by substituents
such as dichloracetyl (U.K. 962,024 and U.K. 1,328,340),
formyl (U.S. 3,641,021), trimethylsilyl or tetrahydro-
pyranyl (U.R. 1,328,340).
There are also provided, according to the present
invention, processes for the preparation of the compounds
of Formula (I), which (a) when X i~ ~
comprlses reacting the compound of the formula
S
H2~~CH- C~ ICH2 ~ - ~
~C N ~ C~C-cH2-s ~ - OH v.
COOH
. Formula (IV)
to produce the compound (in which the hydroxyl group may
i
be protected) having the formula
R-CH-C-NH~ CH CH
~ C ~ 2 ~ ON
COOH
Formula (V)
or the corresponding salt or easily hydrolyzed ester
thereof; and (b) when X is
N=
_~ .,
then reacting the compound of the formula
S
H2N-CH - CH \ CH2
~ C - N \ ~I-CH2-S-~
COOH
Formula (VI)
to produce the compound (in which the hydroxyl gxoup may
be protected) having the formula
R-IH_C_NH fH fH ~H2 /N =l
OB /~ N \ / -CH~-S ~ -N
COOH
Formula tVII)
or the corresponding salt or easily hydrolyzed ester thereof;
or instead of the compounds of Formula (IV~ and of Formula (VI)
with a salt or easily hydrolyzed ester or Schiff base (as
with benzaldehyde) of the compounds of Formula (IV) and of
Formula (VI), said reaction being carried out with an acylat-
ing derivative o~ the acid (in which the hydroxy group may be
protected) having the formula
R-fH-COOH
~B
wherein R is as defined above and wherein B represPnts
hydrogen or the protecting group (that is with that acid or
its reactive derivative substituted at the carbonyl group);
and if such a protecting group is present subsequently sub-
jecting the resulting compound to chemical removal of the
protecting group, that is, subjecting the resulting compound
to elimination reaction of the protecting group.
~o~ 0
The compound of (a) Formula (V) is prepared according
to the present invention by coupl.ing with a particular 3-
thiolated-7-aminocephalosporanic acid designated as Formula
(IV), that is, 7-amino-3-(3-hydroxypyridazin-6-ylthiomethyl)-
3-cephem-4-carboxylic acid, and of (b) Formula (VII) is pre-
pared according to the present invention by coupling with a
particular 3-thiolatPd-7-aminoceph~losporanic acid designated
as Formula (VI), that is, 7-amino-3-(tetrazolo~4,5-b]pyridazin-
6-ylthiomethyl)-3-cephem~4-carboxylic acid; or in each case a
salt or easily hydrolyzed ester or Schiff base as with
benzaldehyde thereof (including, but not limited to, tho~e
of U. S. patent 3,284,451 and U. K. 1,229,453 and any of
the ~ilyl esters described in U. S. patent 3,249,622 for use
with 7-amino-penicillanic acid and used in Great Britian
1,073,530 and particularly the pivaloyloxymethyl, acetoxy-
methyl, methoxymethyl, acetonyl, phenacyl, p-nitrobenzyl
and ~ trichloroethyl estsrs) D-mandelic acid or a
aub~tltuted D-m~ndelic acid a~ de~cribed herein or
thelr runct1~nal equivalent as an acylatln~ agent ~or a
pr~mary amlno groupO Arter ¢oupllngJ any hydro~y
blocking group present 18 remored to give the de~ired
product,
m us, wlt.h respect to said substitut`ed or
unsubstltuted D-mandelic acld to be u~ed to ~.ouple
with compound II, runctional equivalents lnc~lude the
corre~ponding acld anhydrldes, including m~xed
anhydrides and partlcularly the mixed anhydrlde~ pre-
~ared from ~tronger acids such a~ the lower aliph~tic
monoesters o~ carbonic acld~ or alkyl and aryl sulfonic
acld~ and o~ more hindered acids ~uch as diphenylaoetic
acid. A particularly userul anhydride 1~ V-m~ndelio
7~ ~
acid carboxyanh~dride (U.S. 3,167,549) or the corres-
ponding su~stituted D-mandelic acid carboxyanhydride. --
In addition, an acid azide or an ac~ive ester or
thioester (e.g., with p-nitrophenyl, 2,4-dinitro-
phenol, thiophenol, thioacetic acid) may be used
or the free acid itself may be coupled with
compound II after first reacting said free acid with -~
N,N'-dimethylchloroformi~inium chloride [cf. Great
Britian 1,008,170 and ~ovak and Weichet, Experientia
XXI, 6, 360 (1965)] or by the use o~enzymes or of~n
N,N'-carbonyldiimidazole or an N,NI-carbonylditriazole
[cf. South African patent specification 63/2684] or a
carbodiimide reagent ~especially N,N'-dicyclohexylcar-
bodiimide, N,N'-diisopropylcarbodiimide or N-cyclo-
hexyl-N'-(2-morpholinoethyl)carbodiimide; cf. Sheehan
and Hess, J. Amer. Chem. Soc , 77, 1967 (1955)], or
of alkylylamine reagent ]cf. R. Buijle and H. G.
Viehe, A~ . Chem. Internatlonal Edition 3, 582,
(1964)] or of an isoxasolium salt reagent [cf. R. B.
Woodward, R. A. Olofson and H. Mayer, J. Amer. Chem.
$oc. 83, 1010 (1961)], or of a ketenimine reagent
[cf. C.L. Stevens and M. E. Munk, J. Amer. Chem. Soc ,
80, 4065 (1958)] or of hexachlorocyclotriphosphatriazine
or hexabromocyclotriphosphatriazine i(U.S. 3,651,050) or
of diphenylphosphoryl azide ~DPPA; J. ~mer. Chem. Soc.,
94, 6203-6205 (1972)] or of diethylphosphoryl cyanide
[DEPC; Tetrahedron Letters No. 18, pp. 1595-1598 (1973)]
or o~ diphenyl phosphite ~Tetrahedron Letters No. 49,
:
- 8 -
..
,, .. ; .
~LO~'~77~
pp. 5047-5050 (1972)J. Another equlvalent o~ the acid
chloride ls a corresponding azolide, i.e., an amide of
the correspondlng acid whose amLde nitrogen ls a member
o~ a quasiaromatic ~ive membered ring containing at
least two nitrogen atoms, i.e., imidazole, pyrazole,
the triazolesJ benzimidazole, benzotriazole and their
~ubstituted derivatives. As an example of khe general
method for the preparatlon o~ an azolide, N,N'-carbonyl-
dllmldazole is reacted wlth a carboxyllc acid in
equimolar proportions at room temperature in tetra-
hydrofuran~ chlorororm, dimethylrormamide or a ~lmilar
lnert ~olvent to form the carboxyllc acid lmldazollde
in practically quantitative yield with llberatlon of
carbon dioxlde and one mole o~ imidazole. Dicarboxylic
acld~ yield dlmidazolide. The by-product, imldazole~
preclp~tates and may b~ separated and the lmldazolide
laolated, Out this is nok e8~entlal. The methods ~or
carryin~ out these reactlons to produce a cephalo~porin
and the methods used to i~olate the cephalosporin ~o
produced are well known ln the art.
Mentlon wa~ made above Or the u~e Or enz~mes to
couple the ~ree acid with
compound II. Included in the scope Or ~uch processes
are the u3e Or an ester, e.g. the methyl ester, Or
that ~ree acid with enzymes provided by various micro-
organism3, e,g. those descrlbed by T. Takahashi et al~,
J. Amer. Chem. Soc., ~ 111, 4035-4037 (lg72) and by
T. Nara et alDJ J. Antibiotlc~ (Japan) 4(~, 321-323
(1971) and in U.S. 3,682,777~
~,
~05'~770
For the coupling of the substituted or unsub-
stltuted D-mandelic acid ~with or without a protecting
group on the a-hydroxyl) ~s described above with
compound II (or a salt or preferably an easlly
hydrolyzed ester or Schiff base, as with benzaldehyde,
thereor~ it iB also convenient and e~ficient to
utlllze as the coupling agent N ethoxycarbonyl-2-
ethoxy-132-dihydroqulnoline (EEnQ) as described in
J. Amer. Chem. Soc., 90, 823-824 and 1652-1653 (1968)
and U.S. Patent 3,455,929. The reaction ls prererablg
carried out at 30-35 C. in benzene, ethanol or tetra-
hydrofuran using about equimolar quantlties of all
three reagents followed by conventional isolation
and removal by conventional methods o~ any blocklng
groups present.
An additlonal proce~s o~ the present lnventlon
compr~æes the preparation o~ the compound9 of the
present invention by the displacement of the 3-
acetoxy group of a substltuted or unsubstituted
D-mandelamldo-cephalosporanic acid (ln whlch the ~-
hydroxy group is protected or unprotected and so is
the carboxyl grouP) with ~-hydroxy-6-mercapto-
pyridazlne and then removlng the protecting group if
any ls present on the a-hydroxy group or on the
carboxyl group or both, me dlsplacement Or ~uch an
acetoxy group with such a thiol 1~ a well-known
reaction and may be accomplished ln solution as ln
water or aqueou~ acetone at a temperature Or at lea~t
--10--
~ILC)5'~77~ :~
room temperature and preferably within the range of about 50 -
to 100C. in the presence of a mild base such as sodium ;-
bicarbonate, e.g. preferably near neutrality such as at about
pH 6. An excess of the thiol is preferably employed. The
reaction product is isolated by careful acidification of the -
reaction mixture followed by extractio~ with a water-immiscible
organic solvent. Such substituted or unsubstituted D-mandelamido-
cephalosporanic acids are prepared by the procedures described
generally or specifically in J. Med, Chem. 17(1), 34-41 (1974)
and the references cited therein.
In one aspect of this invention there is provided -
a process for making an acid having the D configuration in the
7-sidechain and the formula
101 ~ \ .. ,' '
R-CH-C-NH-CH - CH CH
II ~ 1 2 :~
OH~C _ N \ ~ C-CH2-S-X i
COOH
Formula (I)
wherein X is / N= N
N - N N N ¦ ,~
~/ \ ~ OH or ~ / ~ N, wherein R is a
phenyl group; or
a pharmaceutically acceptable, non-toxic salt thereof, which ;~
comprises (a) when X is N - N :~
~ ~ ~ OH
~ ' ' .
then reacting the compound of the formula
'~
~ ,, `,,
7~
H N-CH --CH / \CH N--N :~
2 ~ ~~ 2 ~\ .i
o// \ ~C--CH2-s - ~ OH .
COO~
Formula (IV)
or (b) when X is ~ N= N
~ N
then reacting the compound of ~he foxmula
~S
H N-CH-- CH ~CH .
,~C N~ ~C-CH2--S ~;~=N
COOH
Formula (VI)
or a salt, easily hydrolyzable estPr or Sc~iff base thereof with :
an acylating derivative of tha acid having the formula :
~ ~' ~
R-CH-COOH ~ .
wherein R is as defined above, and where B represents hydrogen or
a protecting group, to produce the compound of the formula
~S~ ' .,
R-CH-C-NH-CH - CH CH2 .,
OB ~ C - N ~ ~ C-CH2-S-X
COOH .
~ Formula ~V')
: 30 where R, X and B have the meanings above, and if B is a protect-
ing group, subsequen~ly subjecting the compound of Foxmula ~V') i
~ - ll(a~ - ~ ':
1~ .,.
~i~ ' ~.' ,
, .
'7~ :
to an elimination reaction for the protecting group if the
compound of the Formula ~IV) or (VX) is in the form of the
salt, easily hydrolyzable ester or Schiff base, converting
the compound of the Formula (V') to the corresponding compound
of the Formula (I); or if desired, converting the compound of
Formula (V') into the pharmaceutically acceptable non-toxic
salt thereof. ~ -~
In the treatment of bacterial infections in man, the
compounds of this invention are administered parenterally, in
accordance with conventional procedures for antibiotic
administration, in an amount of from about 5 to 200 mg./kg.jday
and pre~erably about 5 to 20 mg./kg./day in divided dosage, e.g.
three to four times a day. They are administered in dosa~e
units containing, for example, 125, 250 or 500 mg. of active
ingredient with suitably physiologically acceptable carriers
or excipients. The dosage units are preferably in the form of `~
liquid preparations such as solutions or suspensions.
.
~,~
~ .
1 .
i
'''
- ll(b) - ~
~s~7~
Startln~ Material~
~-Chloro-6-hydroxv~vridazine
A mixture of 22.47 g. (0~15 mole) of 3y6-dichloro-
pyrldazine and 50 ml. of acetic acid was refluxed rOr
two hours. The reactlon mlxture was cooled and dlluted
with 50 ml. o~ water and then concentrated ~o dryness
under reduced pressure. The re~ldue wa~ cry~talliæed
~rom water to give 15.8 g. (80%) of 3~chloro~6-hydroxy-
pyridazine as colorless prisms which melted at 1~3-7 C.
~lit. 1~8-140 C.). See N. Takabayashl, Yakugaku
Zasshi, 7~, 7~8 (1955).
~-Hvdrox~v~6~mercaDtoP,vrldazine
A mixture of 2 o6 g~ (0~02 mole) of ~chloro-6-
h~droxypyridazine and 5,0 g. (0.07 mole) o~ freshly
prepared pota~ium hydrogen sulfide in ~0 ml. o~ ethanol
wa~ heated at 130-140 C. in a ~ealed tube for 6 hours.
The reactlon mlxture was cooled and dlluted with 200 ml~
of water. Almost all organic solvent was removed by
distlllatlon under reduced pres~ure. The residual
aqueou~ solutlon was acidlfied with dllute hydrochloric
acld to pH 3 and extracted wlth ethyl acetate (6 x 50 ml.),
The combined extracts were evaporated to dryness and
the residue was reprecipitated from 30 ml. o~ ethanol-
llgroln (1:1) to glve 2.2 g. (87~? Or amorphous 3-
hydroxy-6-mercaptopyrldazlne. m.p. 158-159 C.
(llt. 157- 158 C.)0
See J. Drue~ et al,, Helv. Chem. Act~ 121
(1954).
'
i~ SZ7 7
~-Hx-d~o~y--6-~aP~Q~ azi~
To a ~olution of 5.6 g. (0.05 mole) o~ ~J6-dl-
hydroxypyrldazlne ln 150 ml. of pyridlne wa~ added
portlon-wise 2.70 g. ~0.012 mole) of phosphorus
penta~ul~ide wlth vlgorous stlrring under re~luxlng.
The refluxlng was contlnued ~or one hour and then the
reaction-mi~ture wa~ dlluted wlth 200 ml. o~ water
and concentrated to remove the pyridine. The re~ulting
oily re~ldue was suspended in water and extracted with
ethyl acetate. The organic extracts were comblned and
concentrated again to glve oily material which wa~
trlturated with a small amount of water to give 3
hydroxy-6-mercaptopyridazine as a yellow Jolid, Recry~tal~
lization from water af~orded 0.62 g. (12~) of the product
whlch was ldentlcal with that prepared above.
PrePara~
R
¢ _ ~9 ~ HO ~ OH ~ Cl ~ 1
O . - ...
8 ~ 10
N--N
2~]_2_ ~ Cl~ 2
, 11 ., ' , ",
Cl ~ ~ JiS ~ /~~
~2 ~
~05i~77'~ ~
3,6-Dihydrox~pyridazine ~9~
-
To a ~oiling solution of 315 g. ~3 moles) of
hydrazine dihydrochloride in 2 L of water was added
portionwise 295 g. (3 moles) of finely ground maleic
anhydride with stirring. After the addition was
completed the heating was continued for 4 hours and
then allowed to stand overnight in a refrigerator
to give 285 g. ~85%) of 3,6-dihydroxypyridazine as
massive pillars.
m.p. ~ 290C.
3,6-Dichloropyridaz ne (10)
A mixture of 150 g. (1.33 moles) of 3,6-dihydroxy-
pyridazine and 250 g. of phosphorus oxychloride was ;
refluxed for 3 hours under protection from moisture.
The excess of phosphorus oxychloride was removed
under reduced pressure and the dark residue was
poured into one Kg. of crushed ice. The resulting
precipitate was collected by filtration. The second
crop of the product was obtained from the mother
liquor by extraction with five 300 ml. portions
of chloroform followed by treating with 1 g.~of
charcoal and evaporating the solvent. The first and
. . .
second crops were combined, dissolved in 500 ml. of
chloroform and treated again with one g. of charcoal
and concentrated to give 165 g. (83~) of 3,6-di-
chloropyridazine as fine needles melting at 60-61 C. `
(in a sealed tube),
"','
- 14 - ~ ;
~05;~7~
,
3-Chloror~6~hydrazinopyrIda:z~ne ~11~ ;;
A mixture of 40 g. (0.27 mole) of 3,6-dichloro-
pyridazine (10) and 40 ml. of 80~ hydrazine hydrate
in 80 ml. of ethanol was refluxed for one hour. The
reaction mixture was evaporated to dryness and the ',
residue was recrystallized from benzene to give 39 g.
(100~) of 11 melting at 114-115 C. ,~'
6-Chlorotetrazolo[4,5-b]pyridazine (12)
,:- .
To a solution of 25.7 g. (0.174 mole) of 11 in ',
100 ml. of 15% acetic acid was added dropwise a '
soluti~,n of 13.8 g (0.2 mole) of sodium nitrite in
50 ml. of water 'with vigorous stirring at 5~10 C.
Stirring was continued for one hour at the same ''
temperature. The precipitate which separated was ,,
filtered, washed with 20 ml. of water and air-dried '-'
to give 17.02 g. of 12. Additional product was ,~
obtained by evaporation of the filtrate. Total
yield 18.32 g. (64~). M.p. 104-105 C.
6-Mercap o etrazolo[4,5-b]pyridazine (13)
A mixture of 21.3 g. (0.137 mole) of 12 and ~`
20 g. (0.25 mole) of potassium hydrosulfide in 200 ''
ml. of ethanol was refluxed for 2 hours and evaporated
to dryness. The residue was dissolved in 100 ml. of
water and filtered to remove a small amount of
insoluble material. The filtrate was acidified to '~ '
pH 1 with dil. hydrochloric acid to precipitate (13)
as colorless needles which were collected by ~'
filtration, washed with 20 ml. of water and dried.
Yield 9.80 g. (47%). M.p. lQ0-141 C. ~dec.).
,. .. . . .
~0~'~77~
IR; ~ max 2500, 1S40, 1445, 1295, 840 cm
NMR: 2 2 3 7.44 ~1 H, d, 10 Hz, pyridazine-H~,
7.77 ~1 H, d, 10 Hz, pyridazine-_).
Anal. Calcd. For C4H3N5S: C, 31.37; H, 1.97;
N, 45.72; S, 20.94.
Found: C, 31~52; 31.66; H, 1.70; 1.69;!~N,
46.01; 46.01; S, 20.95.
Preparation of 7-amino-3-(tetrazolo[4,5~b]pyridazin-
6-ylthiomethyl)-3-cephem-4-carboxylic acid (3)
S
H2~
O N ~ CH -O-COCH
C2H
7-ACA
.. ~................................................. . . .
-. , ,
N
N- ~ l
N ;
3 ~
~,~
"'.',, :.
~ ,~
~ .i .
2~/~LH2--S~
CO2H
"',,".
''''.
- 16 -
.~ , .
~5;~7 70
7-Amino-3-(tetrazoloI4,5~]pyridazin~6~1thiomethyl~-
3-cephem-4-carboxyl~c acid ~3~.
(i) To a hot solution (50-60 C.) of 9.56 g.
(0.062 mole) of _~-and 10.42 g. (0.124 mole) of sodium
bicarbonate in 300 ml. of water was added carefully
16.86 g. (0.062 mole) of 7-ACA and the mixture was
heated at 80-85C. for 30 minutes. About 7 g. of
sodium bicarbonate was added to the reaction mixture
to dissolve insoluble material. The solution was
treated with active carbon, filtered and the fil~rate
was acidified to p~ 5 with dil. hydrochloric acid.
The precipitate was collected by filtration, washed
with water, air-dried and finally in vacuo on P2O5
to give 14.47 g. (64%) of 7-amino-3-(tetrazole[4,5-b]-
pyrida7in-6-ylthiomethyl)-3-cephem-4-carboxylic acid `
(3). M.p. 248-250 C. (dec.).
(ii) A stirred soltuion of 16.8 g. (0.11 ~ole)
of 13 and 18.48 g. (0.22 mole) of NaHCO3 in 1 L of
0.1 M phosphate buffer (pH 6.4) was heated at 50 C.
and to the solution was added portionwise 30 g.
(0.11 mole) of 70ACA. The mixture was heated at
80 C. for 2.5 hours, during which period insoluble
material still remained. The reaction mixture was
cooled to room temperature and the precipitated
7-amino-3-(tetrazolo[4,5~b]pyridazin-6-ylthiomethyl)-
3 cephem-4-carboxylic acid (3) was collected by
filtration, washed thoroughly with 200 ml. of water
and air-dried.
Additional 3 was obtained from the filtrate and
the washings by acidifying to pH 5 with dil. HCl.
- 17 -
: . . .
~Ll)5Z~7~0
Total yield 32.9 g. (83~). M.P. 245-250 C. (dec.).
IR: KBr 1800, 1615, 1538, 1360 cm 1.
UV: ~ max 3 237 nm ~ 19500), 275 nm (~ 12000),
310 nm (~h) (~ 5700).
NMR ~ D2~K2C3 3.35 (lH, d, 18 Hz, 2-H), 3.76 ( 1 H~
d, 18 Hz, 2-H), 4.00 (1 H, d, 10 Hz,
3-CH2), 4.48 (1 h, d, 10 Hz, 3-CH2),
4~93 (1 H, d, 4 Hz, 6-H), 5.32 t 1 H,
d, 4 Hz, 7-H), 7.46 ( 1 ~, d, 10 Hz,
pyridazine-H), 8.18 (1 H, d, 10 Hz,
pyridazine H).
Anal. Calcd. for C12HllN7O3S2: C, 39.44; H, 3.03; N,
26.83; S, 17.55.
Found: C, 39.19; H, 2.71; N, 26.84; S, 17.35.
6-Ch loro-~-hYdrox~P~ida z ine
A suspenslon o~ 60 g. (0.4 mole) o~ 3,6-dichloro-
pyrldazine in 200 ml. of 10~ hydrochloric acid was
refluxed for 2 hours untll a clear ~olution was obtained.
The clear solution wa s treated with ca . 1.5 g. o~ actlve
carbon and f iltered . The f iltrate was concentrated
under reduced pressure to give 6-chloro-3-hydroxy-
pyridazlne a s colorless needles . Yield 49 .5 g .
( 98%), M . p . 137- 1~9 C .
-18-
~L~)S~770
~-Hydroxv-6-merça~toPyridazi--ne
A mixture of 50 g. (0.38 mole) of 6-chloro-3-
hydroxypyrldazine~ 60 g. (0.8~ mole) o~ potassium
hydrogen sulflde in 250 ml. of ethanol was heated
in a 500 ml. autoclave at 140 C. for 14 hours wlth
magnetic ~tirring. The pressure reached to 15-20
Kg./cm2. The mixture was evaporated to dryne~
and the residue was dissolved in 300 ml. o~ waterO
The aqueous solutlon was acidifled with dll.
hydrochloric acid to pH ~ and extracted wlth ten
200 ml. portions of ethyl acetate. The combined
extract~ were concentrated to give 34.3 g. (70~)
of amorphou3 3-hydroxy-6-mercaptopyridazine.
M.p. 151-152 C.
7-Amino-~-(6-~droxvpyEidazln~ lth
ce~hem-4-carboxYlic acid
A mixture of o.60 g, (0.0047 mole) of ~-hydroxy
6-mercaptopyridazine, 1,27 g. (0.0047 mole) of 7-
aminocephalosporanic acld, 0~78 g. (o.Go94 mole) of
sodlum bicarbonate ln 25 ml. of 0.1 M phosphate
buffer (pH 6.4) wa~ heated at 60 C. for 5 hourg.
The reactlon mixture was filtered to remove a trace
of lnsoluble materlal and the filtrate was ad~usted
to pH 5 with acetic acid to give brown preclpitate~,
whlch were collected by filtration, wa~hed with
water and acetone successively and dried 1~ vacuo
-19~
3L~)5'~7~[)
to glve 7 amlno-3-(6-hydroxypyrldazln-3-ylthio-
methyl)-~-cephem-4-carboxylic acid; 1.0~ g. (71%).
M.p. 290-300 C. (decomp,).
IR: ~ max 1805, 1680, 1650, 1580, 1415 cm 1,
UV ~ 1% NaOH 249 nm (~ 19400).
~D20-~ C0~ 3 22 (lH, dJ 19 Hz), 3.37
ppm
lH, d~ 14 Hz), ~.65 (IH, d, 14 Hz), 3072 (IH, d, 19 Hz),
4.90 (IH, d, 4 Hz), 5.30 (IH, d, 4 Hz), 6.75 (lH, d,
10 Hz), 7.~0 (lH, d, 10 Hz).
Anal Calcd- for C12H12N44S2 ~/2 ~2 9
H, 3.75; N, 16.04; S, 18.36.
Found: C~ 41.45; H, 3.70; N, 15.83; S, 18.03
7-Amlno~ -h~.droxyp~rldaz_n -6-~ iometh~
: ~ephem_4-carboxYli~ acid
: A mixture o~ 141 g. (0.52 mole~) o~ 7-ACA, 92 g.
(1.1 moles) of sodlum bicarhonate and 73 g. (0.57
moles) Or 3-hydroxy-6-mercaptopyridazine ln 1.5 L
of 0.1 M phosphate buffer (pH 6.43 was hea~ed at
60-65 C~ under nitrogen atmosphere for 4 hours. The
hot mixture wa~ treated with 2 g. of charcoal and
rlltered. The filtrate was cooled to room temperature
.
and ad~usted to pH 4.5 with glaclal acetlc acld to
glve the precipitate, which wa~ collected by flltratlon, .
washed wlth one L of acetone and air dried at room
temperature to yield 125 g. (70%) o~ 7-amlno-3-(3-
hydroxypyrldazin -6-ylthiomethyl)-3-cephem-4-~carbox~lic
acid. M.p. 240-250 C, ~dec.).
-20-
1~)5'~:77(~
Plvaloyloxv~neth~Ll Z-Amlno~ (6-h~ydroxvp;~ris~azi~
~lthiometh~ cePhem-4-carboxylate~
Method A. - The tltle compound ls produced by
substltutlng for the 7-amlnocephalo3poranic acld used
immediately above an equimolar weight of pi~aloyloxy-
methyl 7-amlnocephalosporanate hydrochlorlde prepared
accordlng to Example 2 o~ U,K. 1,225,45~ ~rom 7-
amlnocephalo~poranlc acid. German 1,904,585 (Farmdoc
39,445) is equivalent to U~K~ 1~229J453~
Method Bo - The tltle compound is produced by
~ub~tltutlng ~or the 0.025 mole (6.8 g,) 7-amino-
cephalo~poranic acid used in the procedure Or Example
2 Or UoK~ 1,229,453 an equlmolar weight o~ 7-amino-3-
(6-hydroxypyridazln-~-ylthiomethyl3-3-cephem-4-
carboxyllc acid.
The respective acetoxymethylJ methoxymethylJ
acetonyl and phenac~l esters of 7-amino-3-(6-hydroxy-
pyridazin-3-ylthiornethyl)~-cephem-4-carboxylic acid
are prepared by substitutlng in Method B above ror the
chloromethyl pivalate used therein an equlmolar welght
of chloromethyl acetate, chloromethyl methyl ether,
chloroacetone and phenacyl bromide9 respectlvely.
Pivaloyloxymeth~l 7-Amino-3-(tetrazolo[4,5-b]~ridazin-3-
ylthiomethyl)-3-cephem-4-carboxylate.
The methods for producing this compound are the same
as the methods for producing the immediately preceding
one, except that in Method A the reference for substituting
for the 7-ACA, to the description of making 7-amino-3-
(tetrazolo[4,5-b]pyridazin-6-ylthiomethyl)-3-cephem-4-
carboxylic acid; also except that in Method B the substituent
is the same carboxylic acid as mentioned above in this sentence.
-21-
77~
The pre~erred esters of the cephalosporins
of the present invention are the pivaloyoxymethyl,
aceto~ymethyl, methoxymethyl, acetonyl and phenacyl
esters. All are useful intermediates in the pro- ;~
duction of the cephalosporin having a free carboxyl
group and the first three are also of interest
because on oral administration they provide different
rates and amounts of absorption and give differing
concentrations of the active antibacterial agent in ~
blood ~nd tissues. ;
These five esters of 7-aminocephalosporanic ~;
acid are each prepared by known methods. One
excellent procedure is that of U.S. patent 3,284,451
in which sodium cephalothin is esterified by reaction
with the jcorresponding active chloro or bromo
compound (e.g. phenacyl bromide, chloroacetone, :~
chloromethyl ether, pivaloyloxymethyl chloride
[also called chloromethyl pivalate], acetoxymethyl
chloride) and then the thienylacetic acid side-
chain is removed enzymatically as in the same patent
or chemically as in U.S. patent 3,575,970 and in
Journal of Antibiotics, XXIV (11), 7Ç7-773 (1971).
In another good method the triethylamine salt of 7-
aminocephalosporanic acid is reacted directly with
the active halogen compound, as in U.~. 1,229,453.
These esters of 7-aminocephalosporanic acid
., . , ., -.. ..
are then reacted with the nucleophile 3-hydroxy-6-
mercaptopyrid~-ne (which can also be named 6-hydroxy- -
- 22 -
~5'~7~
3-mercaptopyrida~ine or 6~hydroxypyrida~in -3-
thiol) in the same manner as is illustrated
herein for 7-aminocephalosporanic acid itself.
The 3-thiolated ester of 7-aminocephalosporanic
acid is then coupled with the substituted or
unsubstituted D-mandelic acid. Before or
after removal of any blocking group on the ~-
hydroxy group of the 7-sidechain, the ester of
the cephalosporing so obtained is, if not used
per se, converted to its free acid, including -
if desired, any salt by removal of the
esterifying ~group, as by a~ueous or enzymatic
hydrolysis (as with human or animal serum)
or acidic or alkaline hydrolysis or by treat~
, . .
ment with sodium thiophenoxide as t~ught in ;`j ' r
U.S. 3,284,451 and, in the penicillin series,
by Sheehan et al., J. Org. Chem. 29(7), 2006-
2008 (1964).
In another alternative synthesis, the 3-
0 thiolated 7-aminocephalosporanic aacid is prepared
as described herein and then acylated at the
7-amino group and finally esterified, as by
reaction of the appropriate alcohol with the
acid chloride prepared, for example, by reaction
of the final cephalosporin with thion~l chloride
or by other essentially acidic esterification
procedures.
- 23 -
.
1 0~ ~7 7
COCl
C6H5~ cooH ~3> C }~ ~ O
OH . d \ ,~
-~andelic acid_carboxvanhvdride (2)
Pho~gene wa~ bubbled through a solutlon of 2,0 g,
(0.013 mole) o~ D(-)-mandelic acld (1) ln dry tetra-
hydrofuran ~or 30 minutes~ The 30lution wa~ allo~ed
to stand overnight after whlch tlme it was heated
under re~lux for 10 mlnutes, ~vaporation o~ the
nolvent under reduced pressure afforded an olly
re~ldue whlch wa~ ~olldi~ied by trituration with
n-hexane (20 ~l.). The product wa~ colleoted by
~lltration and drled ln vaQ~Q on K0~. Yleld 2.3 g,
o~ D-m~ndellc acid carboxyanhydride.
IR: ~ na~ 18g5, 1875, 1780 om 1,
The prererred and most acti~e compounds o~
the present invention are tho~e havlng the D
conflguration at the ~carbon atom in the 7-side-
chalnJ that i8, those made ~rom D-mandelic acid or
a monosub3tituted D-mandelic acid as lllu~trated here-
ln~ In addltion~ the con~lguration at the two
optically active, a~ymmetrio centers in the ~-lactam
nucleus i3 that round ln cephalo~porin C produced by
~ermentatlon and ln the 7-amlnocephalo~poranic acid
derived there~rom.
-2~-
~Z77~ :
The following example~ are g~:ven in illustration
of, hut not in limitation of, t~e present invention.
All temperatures are in degrees Cent`grade. 7-Amino-
cephalosporanic acid is abbrèviated as 7-ACA; -ACA-
represents the moiety having the structure
-NH-IH IH I 2 -~
~ N ~ C C 2 - ~ .
lOI3, C
COOH O
and thus 7-ACA can be represented as H-ACA-O-C-CH3.
Methyl isobutyl ketone is represented as MIBK.
f'Skellysolve B" is a petroleum ether fraction of
B.P. 60-68 C. consisting essentially of n-hexane. I
i'
Description of the Preferred Embodiments
,.. . . ~
- 25 - j.
,~
, ~
~5f~ 70
E~xamE2;~ 1
CHCON ~S ~ r}
o ~fI <~
C02H
D(-)
4, BB-S283
BB-S28~ 7-~D-a-Hvdroxv-~-~henvlacetamido]-~-~6-
hvdroxy~vridazin-3-~lthlo1nethvl~-~-ce~hem-4-
carb~xy~lic acid (4)
D-(-)-Mandellc acld carboxyanhydride (2) (1.1 g.,
6 mmole~) was added portlonwise to a Ytirred solution
Or 7-amino-3-(6-hydroxypyridazin-3-ylthlomethyl)-3
cephem-4-carboxylic acid (1.02 g., 3 mmoles)
~and NaHC03 (0.84 g., 10 mmoles) in 50% aqueou~
acetone (50 ml.) at O to 5 C. The reaction mlxture `
was stirred at room temperature for one hour and
washed w~th ether (50 ml.). The aqueous layer wa~
acldifled to pH 2 wlth dil. HCl to precipitate~a~
gummy Yub~tance which was collected, washed with
water (10 ml.) and dlssolved in tetrahydrofuran
(THF, 50 ml.). The solution was treated with carbon
and filtered. The filtrate was evaporated under vacuum
below 40 C. to af~ord an oily residue which wa~
solldlfied by trituration with ether (50 ml.),
collected by filtration and washed with ether
(20 ml.) to give o.64 g. (47%) of 7-1D-a-hydroxy-a-
phenylacetamido]-3-(6-hydroxypyrldazin-3-ylthiomethyl)
3~cephem-4-carboxylic acid (4). M.p. 145-152 C. (dec~)
IR ~ KaBX 1780, 1720, 1670~ 1580 cm 1.
U~: ~ la~ C3 252 nm (El%m, 340).
~26-
105~770
Exam~le 2
CHCONH~S --N~ I
OH ,~LN~L~2S~N
D CO~H
4, ~ S261~i
(4)
D-(-~-Mandelic acid carboxyanhydrlde (2) (0.53 g"
3 mmoles) was added portlonwise to a stirred solution
Or 7-amir~o-3-(tetrazolo[4,5-b]pyrldazin-~-ylthlomethyl)- ;
3-cephem-4-carboxylic acld (~) (.73 ~.l 2 mmoles) and
NaHC0~ (o.84 ~., 10 mmoles) in 20 ml, of 50~ aqueous
acetone at 0 C0 The reaction mixture w~8 stirred
at room ternperature for one hour and washed with ether
~30 ml.). me aqueous layer was acidif~ie~ to pH 2
with-dll. XCl and extracted ethyl acetate (2 x S0 mlO )
~reated with a small amount o~ active carbon and
d~ied ~n anhydrous ~odlum ~ul~ate. Removal Or the
solven~ under reduced prescure ar~orded an oily
resldue whlch was triturated wlth ether (50 ml.3 to
glve 7-(D-a-hydroxy-a-p~enylacetami~o)-3-
(tetrazolo~4,5~b~pyrlda~1n~6~ylthlom(3thyl) -7-oeE)hem
-4-carboxylic acid (4). Yleld 0~24 g, (24,g)~ M,p,
i30-135 C0 (dec.~.
.
~5;~Y~O ~:
~" .
IR; ~ mBr 1770, 1720, 1675, 1520 cm 1.
:. .
UV ~ max 243 nm '(~, 17000), 274 nm (sh) (E , 9500),
314 nm (~, 4500).
NMR: ~ ppm 6 4.12 (lH, d, 12 Hz, 3-CH2), 4.70
(lH, d, 12 Hz, 3-CH2), 5.00-6.00 (3H, m, 6~H, 7-H &
C~OH), 7.2 - 7.7 (5H, m, phenyl-H), 7.94 (lH, d, 10 Hz, ;~
pyrida~ine-H), 8.70 (lH, d, 10 Hz~j pyridazine-H).
In vitro Antibacterial Activity of BB-S266
by Tube Dilution Method (Nutrient Broth)O
MIC (mc~.~ml.) ;;
Or~anisms BB-S266
S. aureus Smith 0.1
S. aureus Smith 0.2 ~
+ 50% serum ~ -
S. aureus BX-1633 0.8
S. aureus Russel 0.4 -
St. pyogenes A9604 0.01
Dip. pneumoniae 0.01
Mycobacterium 607 >100
~20 E. coli NIHJ 0.08
E. coli ATCC 8739 1.6
E. coli Juhl A15119 3.1
Kl. pneumoniae A9977 3.1 i~
Kl. pneumoniae A15130 3.1 ',
Pr. mirabilis A9900 6~3 ~;
P~r. morganii A15153 50
.
Sal. enteritidis A9531 0.4
Ser. marcescens A20019 100
Ps. aeruginosa A9843 >100
.~~
- a~ - :
': ' .
:
.... . .~ . , .
105~770 ~:
Example 3
7-[D-~Hydroxy-~phenylacetamido~-3~t6-hydroxy-
pyridazine-3-ylthiometh~ 3-cephem~-4-carboxylic
acid t4) (lot 2)
. '~
A solution of D-(-)-mande]ic acid carboxy-
anhydride (2) (4.5 g., 25 mmoles) in anhydrous
acetone (50 ml.) was added dropw~se to a solution of
7-amino-3-(6-hydroxypyridazin-3-ylthiomethyl)-3-
cephem-4-carboxylic acid (3) (~.0 g., 14.7 mmoles)
and NaHCO3 t4.2 g., 50 mmoles) in 50% aqueous acetone
(200 ml.) with stirring at 0-5 C. After the addition
was completed the stirring was continued for 1 hr. at
ambient temperature. The reaction mixture ~as
extracted with ether (3 x 50 ml.) and the aqueous:llayer
was acidified to pH 2 with dil. ~Cl. The resulting
precipitate was dissolved in THF (200 ml.) and treated
with active carbon (1 g.) The solution was evaporated
under reduced pressure (30 mm. Hg.) below 40 C. The
residue was triturated witheether (200 ml.) to give
2.67 g. of the crude 4. The cEude 4 (2.30 g.) was ~
dissolved in hot THF (40 ml.) and treated with active `
carbon (0.5 g.). The solution was diluted with ether
(300 ml.) and the resulting precipitate was collected
and wa~hed well with ether (100 ml.) Yield 1.6~ g. of
7-[D-~-hydroxy-~-phenylacetamidoJ-3-(6-hydroxypyridazin~
3-ylthiomethyl)-3~cephem-4-carboxylic acid t4) (lot 12)
having m.p. 160-170 C. tdec-)-
IR: ~maxr 1780, 1720, 1670 1580 cm 1.
UV: ~m~K2C3 252 nm (ElCm~ 345).
NMR: ~ppm ~ 5(2H, br-s, 2~CH2), 3.89 (lH,
- 29'-
~s;~
d, 15 Hz, -CH2-S~, 4,17 (lH, d, 15 Hz, -CH2 S~, 5.00
(lH, d, 4 Hz, 6-H), 5~04 (lH, s, ~CH~O~, 5.60 (lH,
d-d, 4 & 9 Hz, 7-H), 6.70 ~lH, d, 10 Hz, pyridazin-H),
7.1 - 7.4 (6H, m, phenyl-H ~ pyridazin-H), 8.6 (lH,
d, 9 Hz, CONH). -
Anal. Calcd. for C20H18N4O6S2
3.82; N, 11.81; S, 13.51.
Found: C, 50.67, 50.39; H, 3.96, 4.27; N, -~
10.69, 10.97; S, 12.85.
Exam~le 4
Sodium 7-[D-~-hydroxy-a-phenylacetamido~-3-(6-
hydroxypyrida~in-3-ylthiomethyl)-3-ceph~m-4-
carboxylate (lot 3)
:
To a solution of 4 (1.0 g., 2.1-mmoles~ in 20 ml. ~-
of THF was added 2.2 ml. of sodium 2-ethylhe~anoate (lM, ,r ,
ethyl acetate solution~ under stirring at room temperature.
The solution was dil~ted with ethyl acetate 1100-ml.) to
afford a precipitate, which was washed well with ~th~l
acetate to give l.C5 g. o i~odium 7-[D-~-hydroxy-a- i~
phenylacetamido]-3-(6-hydroxypyridazin-3-ylthiomethyl)-
3-cephem-4-carboxylate. M.p. 260-270 C. (dec.). ;'
IR: ~mBx 3600-2800, 1760, 1660, 1560, 1580, i
1400 cm 1.
UV: X 2 247 nm (ElCm~ 388).
~2~
NMR: Jppm 3.15 (lH, d, 18 Hz, 2-H), 3.64 (lH,
d, 18 Hz, 2-H), 3.70 (lH, d, 13 Hz, 3-CH2), 4.40
(lH, d, 13 Hz, 3-CH2), 4.90 (lH, d, 4 Hz, 6-H), 5.19
~ S,-CH-o)~ 5.50 (lH, d, 4~Hz, 7-H), 6.80 (lH, d,
10 Hz, pyridazin-H), 7.7 - 7.4 (6H, m, benzen-H &
pyridàzin-H).
- 30 -
~05'~770
Anal. Calcd. rOr C20H17N406S2N~ H2 ~, 46-84;
H, 3.72; N) 10.89; S, 12.46.
Found: C, 47.00, 46.90; H, 3.55, 3.62; N, 10.43,
10.31; S, 12.05, 11.96.
In Yl~Q An~ibacterial Actlvit~ of BB-S283 lot 2
and 3 by Tube Dilution Method (Nutrient Broth).
M ~ c~,/m~J) ___
BB-S283 BB-S283
Or~anlsms ( lot 2~_
S. aureus Smith 0.025 0.05
S. aureu~ Smith 0.1 0.2
~ 50~ serum
S. aureus ~X-1633 o.4 0.8
S~ aureus A15097 103 dil. 100 100
St. pyogene~ A9604 0.02 0.04
Dlp. pneumonlae Ag585 , 0.08 0.04
Enterbacter cloacae A9656 100 50
E. coli NIHJ o.4 0 4
E, coli Juhl!A15119 3.1 6.3
E. coll A9675 3.1 3.1
Kl. pneumoniae A9977 0.8 1.6
Kl. pneumoniae A15130 1.6 3.1
Pr. mirabllis A9900 6 3.1 1.6
Pr. morganii A15153 10 dil. 0.4 0.4
Sal. enteritidis A9531 < 0.2 o.4
Ser. marcescens A20019 10 dil.50 100
P~. aeruginosa A9843 10 dll.> 100 ~ 100
-31-
~L05i~7~
Exam~le 5
Sodium 7~1D-a~hydroxy~-phenylacetamido]~3-tetrazolo-
[4,5-b]pyridazin-6-ylthiomethyl)-3-cephem~4-carboxylate
.,, , _ .~ _ . .
To a solution of 4 (2.1 mmoles) in 20 ml. of
THF is added 2.2 ml. of sodium 2-ethylhexanoate (lM, ethyl
acetate solution under stirring at room temperature.
The solution is diluted with ethyl acetate (100 ml.) to
afford a precipitate, which is washed well with ethyl
acetate to give solid sodium 7-(D-~-hydroxy-a- ~ ;
phenylacetamido)-~-(tetrazolo[4,5-b]pyridazin-6-yl-
thiomethyl)-3-cephem-4-carboxylate.
Example 6
Substitution -for the D-mandelic acid carboxy-
anhydride in the procedure of Example 1 of an equimolar
weight of the carboxyanhydrides prepared in similar
fashion from the monosubstituted D-mandelic acids
D-2-chloro-mandelic acid,
D-3-chloro-mandelic acid,
D-4-chloro-mandelic acid,
D-2-bromo-mandelic acid,
D-3-bromo-mandelic acid,
D-4-bromo-mandelic acid,
D-2-fluoro-mandelic acid,
D-3-fluoro-mandelic acid,
D-4-fluoro-ma~delic acid,
D-2-trifluoromethyl-mandelic acid,
D-3-trifluoromethyl-mandelic acid,
D-4-trifluoromethyl-mandelic acid,
D-3-amino-mandelic acid,
D-3-amino-mandelic acid, ,
D-4-amino-mandelic acid, ~`
D-2-nitro-mandelic acid,
--. , , , , . .. ., ,,., - . . " .. ,
~S'~7`~1
D~3-nitro-mandelic acid,
D-4~nitro~mandelic acid/
D-2-hydroxy-mandelic acid,
D-3-hydroxy-mandelic acid,
D-4-hydroxy-mandelic acid,
D-2-methyl-mandelic acid,
~ D-3-methyl-mandelic acid~
; ~-4-methyl-mandelic acid,
D-2-methoxy-mandelic acid,
D 3-methoxy-mandelic acid .and
D-4-methoxy-mandelic acid respectively produces
7-'(D-2-chloro-mandelamido)-3-(6-hydroxypyridazine-3-
ylthiomethyl)-3-cephem-4-carboxylic acid,
7-(D-3-chloro-mandelamido)-3-(6-hydroxypyridazine-3-
ylthiomethyl)-3-cephem-4-carbaxylic acid, :
7-(D-4-chloro-mandelamido)-3-(6-hydroxypyridazine-3-
ylthiomethyl)-3-cephem-4-carboxylic acid,
7-(D-2-bromo-mandelamido)-3-(6-hydroxypyridazine-3-
ylthiomethyl)-3-cephem-4-carboxylic acid,
7-(D-3-bromo-~andelamido)-3-(6~hydroxypyridazine-3- '~
ylthiomethyl)-3-cephem-4-carboxylic acid,
7-(D-4-bromo-mandelamido)-3-(6-hydroxypyridazine-3
ylthiomethyl)-3-cephem-4-carboxylic acid,
7-(D-2-fluoro-mandelamido)-3-(6-hydroxypyridazine-3- .
ylthiomethyl)-3-cephem-4-carboxylic acid, '.
7-(D-3-fluoro-mandelamido)-3-(6-hydroxypyridazine-3-
ylthiomethyl)-3-cephem-4-carboxylic acid,'
7-(D-4-fluoro-mandelamido)-3-(6-hydroxypyridazine-3-
ylthiomethyl)-3-cephem-4-carboxylic acid, '
7-(D-2-trifluoromethyl-mandelamido~'-3-~6-hydroxypyridazine~
3-ylthiomethyl)-3-cephem-4-carboxylic acid, ~':
7-(D-3-trifluor~methyl-mandelamido)-3-(6-hydroxypyridazine- ; :
- 33 - :
` 1a~5;~:77~
3-ylthiomethyl)-3-cephem-4-carboxylic acid,
7-(D~4-trifluoromethyl~mandelamido~3~(6~hydroxypyridazine-
3~ylthiomethyl~3~cep~em-4-carboxylic acid,
7-(D-2~amino-mandelamido)-3-(6-hydroxypyridazine-3-
.ylthiomethyl)-3-cephem-4-carboxylic acid,
7-(D-3-amino-mandelamido)-3-(6-hydroxypyridazine-3-
ylthiomethyl)-3-cepham-4-carboxylic acid,
7-~D-4-amino-mandelamido)-3-(6-hydroxypyridazine-3-
ylthiomethyl)-3-cephem-4-carboxylic acid,
7-(D-2-nitro-mandelamido)-3-(6-hydroxypyridazine-3-
ylthiomethyl)-3-cephem-4-carboxylic acid,
7-(D-3-nitro-mandelamido)-3-(6-hydroxypyridazine-3-
ylthiomethyl)-3-cephem-4-carboxylic acid,
7-(D-4-nitro-mandelamido)-3-(6-hydroxypyridazine-3-
ylthiomethyl)-3-cephem-4-carboxylic acid,
7-(D-2-hydroxy-mandelamido)-3-(6-hydroxypyridazine-3- `
ylthiomethyl)-3-cephem-4-carboxylic acid,
7-(D-3-hydroxy-mandelamido)-3-(6-hydroxypyri.dazine-3-
ylthiomethyl)-3-cephem-4-carboxylic acid,
7-(D-4-hydroxy-mandelamidoL-3-(6-hydroxypyridazine-3-
ylthiomethyl~-3-cephem-4-carboxylic acid,
7-(D-2-methyl-mandelamido)-3~(6-hydroxypyridazine-3-
ylthiomethyl)-3-cephem-4-carboxylic acid,
7-(D-3-methyl~mandelamido)-3-(6-hydroxypyridazine-3-
ylthiomethyl)-3-cephem-4-carboxylic acid,
7-(D-4-methyl-mandelamido)-3-(6-hydroxypyridazine-3-
ylthiomethyl)-3-cephem-4-carboxylic acid, ;
7~(D-2-methoxy-mandelamido)-3-(6-hydroxypyridazine-3-
ylthiomethyl)-3-cephem-4-carboxylic acid, ;
7-tD-3-methoxy-mandelamido)-3-(6-hydroxypyridazine-3-
ylthiomethyl)-3-cephem-4-carboxylic a~id`and
7-(D-4-methoxy-mandelamido)-3-(6-hydroxypyridazine-3- -
ylthiomethyl)-3-cephem-4-carboxylic acid respectively.
- 34 -
~- . . . ~ ~ , . . . . .
:'.. '- .', ' : ' ' : ' '',. ':
5'~7'-~0
Exam~e 7
Substitution for the D~mandelic acid
carboxyanhydride in the procedure of Example 1 of an
equimolar weight of the carboxyanhydride prepared in
similar fashion from D-2-thiopheneglycolic acid and ;~
D-3-thiopheneglycolic acid respectively produces
7-(D-~-hydroxy-2-thienylacetamido~+3-(6-hydroxypyridazine-
3-ylthiomethyl)-3-cephem-4-carboxylic acid and 7-(D-~-
hydroxy-3-thienylacetamido)-3-(6-hydroxypyridazine-3-
ylthiomethyl)-3-cephem-4-carboxylic acid respectively.
Example 8
Substitution for the D-mandelic acid
carboxyanhydride in the procedure of Example 2 of an
equimolar weight of the carboxyanhydride prepared in
similar fashion from D-2-thiopheneglycolic acid and -
D-3-thiopheneglycolic acid respectively produces
7-(D-~-hydroxy-2-th-~enylacetamido)-3-(~*etrazol~f~5r~b]
pyridazin~6~ h~ome.th~ ce~ éin~l4~,c~bo~yl~:caci~d~
and 7-~D-a-hydroxy-3-thienylacetamido)-3-(tetrazolo-
[4,5-b]pyridazin-6-ylthiomethyl)-3-cephem-4-carboxylic
acid respectively.
Exam~le ~
7-(D~a-Hydroxy-a-phenylacetamido)-3-(6-hydroxy- ~,-
pyridazin-3-ylthiomethyl)-3- ~ rboxylic acid
prepared from 7-D-mandelamidocephalo-s~oranic ac~d.
0.27 Mole of sodium 7-D-mandelamidocephalo-
sporanate i9 suspended in 1000 ml. of 0.1 M phosphate
buffer of pH 6.4 and there is added 0.31 moles of
3-hydroxy-6-mercaptopyridazine. The solution is heated
,i
at 55 C. under a nitrogen atmosphere for five hours.
After one hour the pH is adjusted to 6.4 by addition
of a small amount of 40~ H3PO4. At the end of the fi~e
:, ',:
~ 35 _
,,
s~
hour heatin~ period the ~olut~ion i~ cooled to 23 C.
and the pH adjusted to 2 ~y the add~tion of 3 N HCl
under a layer of ethyl acetate. The product is extracted
into ethyl acetate and stirred for 15 min. at 23 C. with
2 g. of ("Darco KB'') decolorizing charcoal. The mixture
is then filtered through a pad of diatomaceous earth ("Celite'')
and the ethyl acetate removed from the filtrate under vacuum.
The residue is triturated to a solid with diethyl ~ther,
collected by filtration and dried over P2O5 under vacuum
to yield solid 7-(D-~-hydroxy-~-phenylacetamido)-3-(6- ,,
hydroxypyridazin-3-ylthiomethyl)-3-cephem-4-carboxylic acid.
Example 10
. . .
7-,~D-~-Hydroxy-~-phenylacetamido)-3-(tQ~razolo-
[4~5-b]pyrida~in-6-ylthiomethyl ? -3-cephem-4-carboxylic
acid ~repared from 7-D~mandelamidoce~halosporanic acid,. '
0.27 Mole of sodium 7-D~mandelamidocephalo-
sporanate i5 suspended in 1000 ml. of 0.1 M phosphate
buffer of pH 6.4 and there is added 0.31 moles of 6-
mercaptotetrazolo[4,5-b]pyridazine. The solution is heated ~,~
, 20 at 55 C. under a nitrogen atmoshere for five hou~s. After '
one hour the pH is adjusted to 6.4 by addition of a small ,
amount of 40~ H3PO4. At the end of the five hour heating
period the solution is cooled to 23 C. and the pH adjusted -
to 2 by the addition of 3`~,N HCl under a layer of ethyl -1
acetate. The product is extracted into ethyl acetate and
stirred for 15 min. at 23 C. with 2 g. of tllDarco KB")
decolorizing charcoal. The mixture is then filtered through
a pad of diatomaceous earth ("Celite") and ~h~ ethyl acetate
removed from the filtrate under vacuum. The residue is tri-
turated to a sol~d with diethyl ether, collected by filtration
and dried over P205 under vacuum to yield solid 7-(D~
hydroxy-~-phenylacetamido)-3-t~etrazolo~4,5-b]pyridazin-6-
ylthiomethyl)-3-cephem-4-carboxylic acid.
* Trade Marks 36 -
