Note: Descriptions are shown in the official language in which they were submitted.
o
This invention relates to a no-vel cephalosporin
E~ derivati~e and a method for producing the same.
More particu'arly, thi~ invention pertains to
pivaloyloxyme-thyl 7~-[2-(2-aminothiazol-4-yl)acetamido]-
3-[[~1-(2-dimethylaminoethyl)-lH-tetrazol-5-yl]thio~methyl]-
ceph-3-em-4-carboxylate having the formula (I):
H2N ~3
N - - Cl2C,ONE ~ 9 ~ - N
CH2CH2N~
COOCH20~C~a(CH3)3
and its acid addition salts, and also to a method for the
production therefor.
.
~ :L - ~
Extensive studies made by the present inventors led to
the followin~ findin~s, namely the above-mentioned cephalo-
sporin derivative (hereinafter sometimes referred to as
"ester compound") and its acid addi-tion salts are readily
absorbed into body through gastro~intestinal tract and the~
are hydrolyzed under the influence of the enzymes present in
the host to the corresponding free carbo~ylic acid compound
(hereinafter sometimes referred to as "non-ester compound"),
which has antibacterial activity against both gram-positive
and gram-negative bacteria including such ones as having
acquired resistance against so far known cephalosporanic
antibiotics, thereby the concentration of the non-ester com-
pound in blood reaches a high level enough to show the thera~
peutic effect, and therefore the ester compound and its acid
lS addition salts are useful as an orally administrable anti-
biotics having a very broad anti-microbial spectrum.
The above-mentioned acid addition salts are formed at
the aminothiazol group and/or dimethylamino group of the
ester compound, thereby the properties such as water-solubility,
absorption efficiency and stability/ of the ester compound may
be further improved. As preferable acids suitable for forming
such acid addition salts, there may be mentioned well-known
acids for forming pharmaceutically acceptable salts
in the field of penicillins and cephalosporins. Namely, they
include mineral acids such as hydrochloric acid, sulfuric acid,
phosphoric acid and the like, and organic acids such as maleic
acid, acetic acid, citric acid, succinic acid, tartaric acid,
benzoic acid, fumaric acid, malonic acid, mandelic acid,
- 2 -
5%~
ascorbic acid arld the like.
The ester compound (I) of the present invention can be
produced ~y, for example, the following methods:
Method 1
The compound (I~ can be produced by esterifyIng the
non~ester compound having the formula (II):
N 1 CH2CON~ C~2S ~N N
CH2CH2N ~
COOH CH3
or its salt with the compound of the ~ormula (III):
HOCH20~ C(CH~)3 (III)
or its reactive deri~ative.
As said reactive derivative, there may be used per se
known derivatives, especially, esterification by means of a
compound of the formula: XCH20l~C(CH3)3 ~wherein X is a
halogen atom) is prefera~le. The salts of the non-ester com-
pound may be exemplified by acid addition salts with mineral
acids such as hydrochloric acid, sulfuric acid or nitric acid,
or organic acids such as oxalic acid or p-toluene sulfonic
acid, or salts with alkali metals such as sodium or potassium,
alkali earth metals such as calcium or magnesium or bases such
as triethyl amine, ~rimethylamine, pyridine, colidine or
.
~ - 3 -
~7~
lutidine. As halogen atom, any o~ chlorine, bromine and iodine
may be used. From standpoint o~ ~eactivity iodine and bromine
are preferable, the Eormer beiny most preferred.
The esterificatlon reaction is generally ~arried out in
a solvent inert to the reaction~ As such solvents, there may
be mentioned amides, ketones, nitriles, liquefied sulfurous
acids and so on. Typical examples of these solvents are
acetonitrile, N,N-dimethylformamide (DMF), N,N-dimethylacet-
amide (DMA), dichloromethane, chloroform, dimethylsulfoxide
(DMSO), diethylether, dioxane, tetrahydrofuran (THF), acetone,
methyl ethyl ketone, etc. ~mong them, DMF, acetone, aceto-
nitrile and liquefied sulfurous acid anhydrid~ are particularly
preferred. The reaction may preferably be carried out in the
presence of a base. Any base capable of acting as deacidation
agent can be used. For example, there may be used organic
amines such as dicyclohexylamine, N-ethyl aniline, morpholine,
N,N-diethyl aniline, N-methyl morpholine, pyridine, triethyl-
amine, etc. or inorganic bases such as sodium hydrogen carbonate,
lithium carbonate or potassium hydrogen carbonate. The amount
of the base to be used i5 desirably not less than equimolar to
the non-ester compound or its salt. The esterification reaction
is carried out generally at a temperature in the range from
-20 to 20C. When liquefied sulfurous acid anhydride is used
as a solvent, the reaction is carried out preferably at a
temperature near the boiling point of this solvent (-lO~C),
namely at -10 to -20QC. The reaction time may differ, depend-
ing on the reactants and the solvent employed, but is generally
in the range from about 10 minutes to about 6 hours.
~ - 4 -
~7~
Method 2
According to another process, the compound (I) can
be prepared by first reacting a compound of the formula
(II'):
A ~ 1 N- ~ ~
N ~ 2 N CH3 (II')
COOH CH2CH2N - CH3
wherein A is amino group or an acylamino group e~cluding
(2-aminothiazol-4-yl)acetamino group, or its sa].t, with the
compound of the formula (III) to carry out the reaction in
the same manner as the esterification reaction as previously
described~ and then, in case when A is an acylamino group,
deacylating the esterified product by a conventional method
(as disclosed by Journal of Medicinal Chemistry, Vol. 18,
p.992, 1975) to derive the compound of the formula (IV);
H2N~
N ~ CH2S ~ ,N CH (IV)
¦ CH2CH2N -CH3
COOCH20CC(CH3)~
and further reacting tha compound (IV) with the compound.
of the formula (V):
~2~ ~ S
11 CH2COOH (V)
or its reactive derivative, thereby to effect acylation of
the amino group at 7-position of the compound (IV).
_ 5 _
When A in the above Formula (II~) is an acylamino group,
the acylamino g~oup may be arly one known ~er se in the art of
cephalosporin compou~ds Examples of the acylamino group are
acetylamino, benzoylamino, phenylacetylamino, thienylacetyl-
amino, phenyloxylacetylamino, 5-amino-5-carbox~valerylamino,
and so on. When A is amino group or an amino-containing acyl-
amino group, the amino group is preferably protected during the
reaction. As the protective group of the amino group, there
may be used per se known protective groups of amino groups,
including t-butoxycarbonyl, carbobenzyloxy, 2-hydroxy-1-
naphthocarbonyl, trichloroethoxycarbonyl, 2-ethoxycarbonyl-1-
methylvinyl and 2-methoxycarbonyl-1-methylvinyl. The amino
group of the compound (V) may also be protected during the
reaction. This protective group may be the same as that of the
amino group as A in the compound (II'). The compound (V) can
be used either in free form or in the form of a reactive
derivative thereof. Namely, it can be provided for acylation
reaction either as free acid, as a salt with sodium, potassium,
calcium, trimethylamine, pyridine, etc., or as a reactive
derivative thereof such as acid halide, acid anhydride, mixed
acid anhydride, active amide, ester, etc. As an active ester,
there may be used p-nitrophenyl ester, 2,4-dinitrophenyl ester,
pentachlorophenyl ester, N-hydroxyphthalimide ester, etc. As
a mixed acid anhydride, there may be used a mixed acid anhydride
with mono-ester carbonates such as mono-methyl carbonate, mono-
isobutyl carbonate or a mixed acids anhydride with lower
alkanoic acids which may be substituted by halogens such as
pivalic acid or trichloroacetic acid.
~ - 6 -
5~
When the compound ~V) is used in its free form or an
acid form, there may be used a condensing agent as exemplified
by N,N'-disubstitl~ted carbo~iimides such as N,N~-dicyclohexyl-
carbodiimideJ azolide compouncls such as N,N'-carbonylimidazole
or N,N'-thionyldiimiclazole; dehydrating agents such as N-
ethoxycarbonyl~2-ethoxy~1,2-dihydroquinoline, phosphorus
oxychloride, alkoxy acetylene (e.g. ethoxy acetylene); and so
on. When these condensing agents are used, the reaction is
considered to proceed via a reactive derivative of the carbox-
ylic acid.
The present reaction can usually be practiced advanta-
geously and smoothly in a solvent. As a solvent, there may be
used any of the solvents in general which do not interfere
with the reaction, or a mixture thereof, including water,
acetone, diisobutyl ketone tetrahydrofuran, ethyl acetate,
dioxane, acetonitrile, chloroform, dichloromethane, dichloro-
ethylene, pyridine, dimethylaniline, dimethylformamide,
dimethylacetamide, dimethylsulfoxide, and so on. The reaction
temperature is not particularly limited, but the reaction is
conducted usually under cooling or at room temperature. When
the reaction proceeds with liberation of acids, bases are
allowed to coexist, if desired~ As such bases, there may
generally be used aliphatic, aromatic or heterocyclic nitrogen
bases or alkali metal carbonate or bicarbonates, for example,
triethylamine, N,N'dimethylaniline, N~ethyl morpholine,
pyridine, collidine, 2,6-lutidine, sodium carbonate, potassium
carbonate, sodium hydrogen carbonate and potassium hydrogen
carbonate. When the acylation reaction proceeds preponderantly
by way of dehydrating reaction, it is preferred to exclude
~ - 7 -
water from the solvent. In some cases, the reaction may be
operated in an inert ~as such a$ nitrogen so as to be kept
away ~rom moisture.
M~
According to still another process, the compound
(I) can be produced by reacting the compound (IV) a~
mentioned above with 4-halo~eno-3-oxobutyrylhalogenide,
which is obtained by reacting diketene with equimolar
amount of halogen (e.g. chlorine, bromine or iodine), to
obtain the compound (VI) of the ~or~ula:
0 ~ 2 N CH (VI)
¦ CH2CH2N ~ 3
COOCH20CC(CH3)3
O
wherein Y represents a halogen atom (e.g. chlorine, bromine, or
iodine atom), and then reacting the compound (VI) with thiourea.
In the reaction between the compound (VI) and thiourea, thio-
urea can be provided for the reaction as it is or as a salt
with an alkali metal such as lithium, sodium or potassium or
an ammonium salt. The reaction is usually carried out by
mixing equimolar amounts of both compounds and 1 to 2 equiva-
lent of a base. As a solvent suitable for this reaction, there
may be mentioned ~ater, methanol, ethanol, acetone, dioxane,
acetonitrile, chIoroform, ethylene chloride, tetrahydrofuran,
ethyl acetate~ dimethylformamidel dimethy~acetamide and other
common organic solvents which do not interfere with the
8 -
7~ ~
reaction. ~mong them, a hydroph:ilic solvent can be used in ad-
mixture with water. As a base to be used, there may be mention-
ed alkali me-~al hyclroxides such as sodilIm hydroxide, potassium
hydroxide, e-tc.; alkali metal carbonates such as sodium
carbonate, potassium carbonate, etc.; alkali met~l hydrogen
carbonate such as sodium hydrogen carbonate; and organic
tertiary amines such as trimethylamine, triethylamine, pyridine,
etc. The base ls used for the purpose of neutralizing carboxyl
groups of cephalosporin and hydrohalogenic acid eliminated by
the reaction. Thus, it is used usually in about 2 oquivalent
amount as free thiourea and in about 1 equivalent amount as an
alkali metal salt, respectively. The reaction temperature is
not particularly limited, but it is generally preferred to
carry out the reaction under cooling. The reaction generally
takes place rapidly and is completed usually within 10 minutes,
but it may sometimes take more than 30 minutes before comple-
tion of the reaction.
When ~ -isomer is co-present with the objective ester
compound (I) in the reaction product obtained by these reac-
tions, it may be isomerized, if necessary, by per se knownmethod, for example, the method as disclosed in Journal of
Medicinal Chemistry, Vol. 18, p. 986, 1975, into ~3~isomer or
converted to ~3-isomer by leading to corresponding S-oxide
thereof, followed by reduction of the a3-isomer to be reversed
to the ester compound (I) by conventional methods.
The objective compound (I1 can be isolated and purified
according to conventional methods such as solvent extraction,
pH adjustment, phase transfer, crystallization, recrystallization,
chromatography, etc.
_ g _
When free ester compol~d is obtained by the method as
described above, it may be converted to a pharmaceutically
acceptable acid addition salt or a:Lternatively, when the com-
pound is obtained in the form of a salt, it may be converted
to a free form, by per se known methods, respectively.
Further, the aminothiazole group in the above compounds may
sometimes be present as tautomer, i.e. iminothiazolin group.
The thus produced ester compound (I) or a pharmaceutic-
ally acceptable acid addition salt thereof is diluted with
diluents for medicines by per se known methods to prepare an
orally administrable cephalosporin medicament of the present
invention. Dilution may be carried out by conventional
methods known per se such as mixing. Examples of suitable
diluents are starch, lacto$e, sucrose, calcium carbonate,
calcium phosphate and the like. There may also be compounded,
if desired, with other additives. Preferable additives are,
for example, binders (e.g. starch, gum arabic, carboxymethyl
cellulose, hydroxypropyl cellulose or crystalline cellulose),
lubricants (e.g. magnesium stearate or talc) or disintegrators
(e.g. carboxymethyl calcium or talc). After various components
are mixed, the mixture is made into various forms suitable
for oral administration by per se known methods such as
capsules7 dispersants, fine particles, granules or dry syrup.
The orally administrable cephalosporin preparations
of the present invention can be used as an antibiotic having
broad spectrum for oral administration for treatment of
in~ectious diseases caused by ~arious gram-positive and gram-
-- 10 --
75,2~
nega-tive bacteria. When the preparation oE the present inven-
tion is orally administered the es-ter compound or a salt
thereof is absorbed from gastro-intestinal trac-t and hydrolyzed
immediately by enzymes in a body to be conver-ted to corres-
ponding non-ester compound or a sal-t thereof. The non-ester
compound and its salts have especially excellent anti-microbial
activity. Namely, said non-ester compound and its salt
exhibit excellent anti-microbial activities against gram-
positive bacteria such as S*aphylococcus aureus, and gram-
negative bacteria such as Escherichia coli, Klebsiellapneumoniae, Proteus vulgaris, Proteus mirabilis and Proteus
morganii. Further, the ester compound and its salts are low
in toxicity. Accordingly, the cephalosporin preparations
of the present invention can be used as effective orally
administrable antibiotics for treatment of various diseases
caused by these bacteria.
More specifically, the cephalosporin preparation of
the present invention can be used for treatment of, for
example, pyogenic diseases, biliary tract, respiratory and
urinary tract infections caused by the aforesaid bacteria
in human beings and other warm-blooded animals (e.y. rat,
mouse, rabbit, horse, dog, monkey, etc.). The cephalosporin
preparation of the present invention may be administered
by oral route, for example, in case of pyogenic dieases
in adult human, at a dosage of about 3 to 4 times administra-
tions in a dose of about 1 to 20 mg/kg-body weight per every
administration, calculated as non-ester compound.
, .
-
The present inven-tion is described in further detail
by referring to the following Examples, by which ~he present
invention is not limited.
Example l
Preparation of pivaloyloxymethyl 7~-[2-(2-aminothiazol-
4-yl)acetamido]-3-[[[1-(2-dimethylaminoethyl)-lH-tetrazol-5-
yl]thio]methyl]-ceph-3-em-4-carhoxylate:
In 60 ml of dimethylformamide, there is dissolved 5.9
g of potassium 7~-[2-(2-aminothiazol-4-yl)acetamido~-3-[[[1-
(2-dimethylaminoethyl)-lH tetrazol-5-yl]thio]methyl]-ceph~3-
em-4-carboxylate. While stirring the mixture under ice-
cooling, a solution of 2.4 ~ pivaloyloxymethyl iodide in
5 ml dimethylformamide is added dropwise for 10 minutes.
After the mixture is stirred for additional 20 minutes,
the reaction mixture is mixed with ethyl acetate (1 ~) and
washed with water (150 ml x 4), then with a saturated aqueous
sodium chloride solution (100 ml x 2), and dried over
anhydrous sodium sulfate. This solution is concentrated
under reduced pressure to give white powders, which are
washed out with ethyl ether, followed by suction filtration,
and dried under reduced pressure to obtain the objective
compound. ~ield 4.2 g, m.p. 84-87C.
IR(KBr cm ): 1780, 1735
NMR(90 MHz, in d6-DMSO, ~): 1.17(s,(CH3)3C),
2.16(s,N(CH3)2, 2.65(t, J=6Hz, NCH2), 3.35(s,2~CH2),
4.18 and 4.46(ABq, J=15Hz, 3-CH2), 4.33(t, J- 6Hz, NCH2),
5.06 (d, J=5Hz, 6-H), 5.6-5.9 (m,7-H and, O-CH2-O),
6.20(s, thiazole-5H), 6.76(broad s, -NH2), 8.82(d,
J=7.5Hz, CONH)
- 12 -
.
z~
Elemental analy,si~
Calculated for C24H33N96s3 2~2
C, A4.Al; H, 9.24; N, 19.42
Found: C, 44.53, E-l, 5.11; N, lg.l6
This product (1.0 g~ is dissolved in 100 ml of
ethyl acetate and mixed with ethereal dry hydrochloric
acid to precipitate white powders. After suction
filtration under reduced pressure, the powders are dried
to obtain di-hydrochloride of the title compound.
~ield: 630 mg
IR(KBr, cm 1): 1770, 1735
NMR(~0 MHæ, in D2O, ~): 1.21(s,(CH3)3C),
3.08(s,N(CH3)2), 3.7-4.5(m,N(CH2) and 2-C~2), 4.97(t,
J=7Hz,CH2N - CH3)' 5.20(d, ~=4.5Hz,6H), 5.68-5.85(m,
15 O-CH2-O), 5.93(d,J=4.5Hz), 6.71(s,thiazole-5H)
Elemental analysis.
24 33 9 6 3 2
C, 39.45; H, 5.06; N, 17.26
Found: C, 39.50; H, 5.28; N, 16.46
Example 2
Various pharmaceutical compositions are prepared
according to the following procedures.
(1) The principal medicine and lactose are
previously mixed and to the resultant mixture is added
a 10~ aqueous hydroxypropyl cellulose solution. The
mixture is kneaded, then dried and crushed to prepare
particles. ~he particles are mixed with magnesium
stearate previously dispersed in starch and molded into
tablets having the composition as below.
,~
r-~
,-3
Pivaloyloxymethyl 7~-[2-(2-amino-
thiazol-4-yl)acetamido]-3-[~[1-
(2-dimethylaminoethyl~ 1-tetrazol-
5-yl]thiolmethyl]-ceph-3-em-4-
car~oxy]ate 76.0 mg
Lactose 16.0 mg
Starch 5.0 mg
Hydroxypropyl cellulose 2.7 mg
Magnesium ~tearate 0.3 mg
100 mg/tablet
Similar tablets are prepared by repeating the above
procedure except that the above principal medicine is re-
placed by its hydrochloric acid salt.
~2) a) A mixture of a part of starch and magnesium
stearate is blended with the principal medicine and residual
starch. The resultant blend is made into capsules by
conventional encapsulation method, the composition of which
is as below.
Pivaloyloxymethyl 7~-~2-(2-aminothiazol-
4-yl)acetamido]-3--[~ (2-dimethylamino-
ethyl)-lH-tetrazol-5-yl~thio]methyl]-
ceph-3-em-4-carboxylate 76 mg
Starch 22 mg
Magnesium stearate 2 mg
100 mg/capsule
bl By the same method as descrihed in a),
capsules are prepared from the following composition:
J r ~
L IL O ~d,r~'
Pivaloyloxymethyl 7~-[2-(2-amino-
thiazol-4-yl)acetamido]-3-3-~[[1-
(2-dimethylaminoethyl)-lH--
tetrazol-5-yl]thiolmethyl]-ceph-3-
em-4-carboxylate dihydrochloride 84.7 mg
Starch 37~3 mg
Magnesium stearate 3 mg
125 mg/capsule
(3) The principal medicine, starch and lactose are
prevîously mixed and then kneaded with a 10~ aqueous hydroxy-
pyropyl cellulose solution, followed by drying and crushing
to prepare particles, which are then separated by a sieve
into fine particles with sizes of 32 to 150 mesh, whose
composition is as below.
Pivaloyloxymethyl 7~-[2-(2~amino-
thiazol-4-yl)acetamido~-3-[[[1-
(2-dimethylaminoethyl)-lH-
tetrazol-5-yl]thio]methyl]-ceph-
3-em-4-carboxylate 76 mg
Lactose 12 mg
Starch 9 mg
Hydroxypropyl cellulose 3 mg
100 mg
Similar fine particles are obtained by using corres-
ponding hydrochloride in place of the principal medicine in
the above composition.
(4) Particles are prepared by the same method as
in (31 and separated by a sieve into granules with sizes
of 12 to 48 mesh, whose composition is as below.
- 15 -
Pivaloyloxymethyl 7R-[2-(2-amino-
thiazol-4-yl)ace-tamidol-3-~[[1~
(2-dimethylaminoe-thyl)~ tetrazol-
5-yl]thio]methyll-ceph-3-em-4-
carboxylate 76 rng
Lactose 16 mg
Starch 4 mg
Hydroxypropyl cellulose 4 mg
100 mg
(5) The principal medicine, sucrose, citric acid
anhydride and 0.5 ml of water are kneaded, followed by
drying and crushing, to prepare a dry syrup having the
following composition.
Pivaloyloxymethyl 7~-[2-(2-amino-
thiazol-4-yl)acetamido]-3-[[[1-
(2-dimethylaminoethyl)-lH-
tetrazol-5~yl]thio]methyl~-ceph-
3-em-4-carboxyla~e 76 mg
Sucrose 90 mg
Citric acid anhydride 10 mg
176 mg
In place of the above principal medicine, corresponding
hydrochloride is used to prepare similar dry syrup.
Example 3
Acute toxicity test
The cephalosporin preparations of the present inven-
tion are administered orally to mouse and rat to examine
acute toxicity to give the following results.
Subject: 5 male mice (ICR strain 5 weeks old)
5 male rats (SD strain 5 weeks old)
Method: The ester compound obtained in Example 1
is made into a 5% gum arabic suspension
- 16 -
111 ~r
and administere~ orally to the animals in
doses oE 0.5 to 3.0 g/ky. The amount of
liquid is 0.2 ml/10 q~body weight.
Observation Term: 7 days
Result: LD50(g/kg)
Mouse >3.0
Rat >3.0
Example 4
Oral administration test
The cephalosporin preparations of the present
invention are administered orally ~o human being to give
the results of ~lood level of the non-ester compound and
urinary recovery as shown below.
Subject: 3 Healthy adult human volunteers
Dose: The ester compound obtained in Example 1:
152 mg, or dihydrochloride of the ester
compound: 169.4 mg, each corresponding
to 125 mg of the non-ester compound.
Method of administration: Two capsules of ~2) a)
or b) in Example 2 are orally adminis-
tered.
Quantitative method: Proteus mirabilis is used
.
as the bacteria to be examined and
quantitative analysis is conducted by
- cylinder-plate assay.
- 17 -
~loo~-le_el (mcq/ml: avera~e o:E -thre _sam_les)
_ _ Time (hours) __
~.~ 1 2 4
Capsule of
Example 2 (2)
a) (ester
compound) 1.204.0 2.1 0.30
Intramuscular
administration
of non-ester
compound
(reference) *1 6.33 4.80 2.1 0.43
*l) dose: 125 mg
Urinary recovery (%)
0 - 6 hours: average of three samples
Capsule of Exampl~
2 (2) a) (ester
compound) . 51.5
Capsule of Example
2 (2) b) (ester
compound dihydro-
chloride) 50.5
Intramuscular
administration of
non-ester
compound
(reference) *168.0
*l) dose: 125 mg
The ester compound after being absorbed is
hydrolyzed at 4-posi-tion carboxylic acid e.ster and found
to exist in either blood or urine as non-ester compound
of the corresponding 4-position carboxylic acid of the
cephalosporin compound.
- 18 -
~7
.x;:~mp:L~? 5
1) E'reparation of pivaLoy~LoxyrQe-thyl 7~-arnino-3-
r[ [1-(2-di~ne-thy:Larrli.noe thy.l.)-l.I-I-tet:r,lzol~5-;yl]thio]methyl]
ceph-3-em-4-carboxylate ~3ihydrochlorlde[foYmula (IV)3
To a mix-ture of 7~-atn~no-3-~1-(2-dirnethy]amino-
e-thyl)-lH-tetrazol~5-yl~-thio]methyl3ceph-3-em~4-carboxylic
acid hydrochlori~e (4.22 g) and potasslum acetate (1.67 g)
in dimethylacetamide (60 ml) is added dropwise pivaloyloxy-
methyl iodide (2.42 g) at oa with stirring, and then the
mixture is stirred for 5 minutes at 0C. To the mixture
was added a mixture of water (60 ml) and dichloromethane
(60 ml), and adjusted to pH 2 with N-HCl. ~he aqueous layer
is separated, and this layer is adjusted to pH 6 with aqueo~s
sodium hydrogen carbonate and extracted with dichloromethane
(60 ml x 2). On the organic layer is layered water and
E~ adjusted to pH 2 with 4N-Hal. After removing dichloromethane
under reduced pressure, the solution is lypophilized -to give
the objective compound (IV3. Yield 2.02 g.
IR(KBr cm 1): 1785, 1750 ~ ~
l~MR(in D20,~): 1.23(s, (CH3)~), 3.1(s, ~(CH3j2), 3.91 (m,
CE2CO and a3-H), 3.96(t, J=7Hz, CH2N), 4.36(b-s, C3-H)~,
5.01(t, J=7 Hz, CH2N), 5.20(d, J=4.5 Hz, C6-E), 5.~6(d,
J=4.5 Hz, C7-H) and 5.86(ABq, J=6Hz~ -O-CH2-0-)
b) Preparation o~ pivaloyloxymethyl 7~-[2-(2-
aminothiazol-4-yl)acetamido]-3-[[[1-(2~dimethylaminoethylj-
lH-tetrazol-5-yl3thio]methyl]ceph-3-em-4-carboxylate.
~ he above compound (IV) (1.15 g) is stirred with
sodlum hydrogen carbonate (0.~4 g) in a mixture o~ dichloro-
methane (30 ml ) and water (30 ml). ~he organic layer is
separated, drled over anhydrous calcium chloride.
.
-- 19 --
.
'' .
rl'~l~' dr~ iri~ r~ f i l t~-~r~(l o:ff, arl~ -t;he :fi~.tratf~ i3 added
:ir~ 1 so~Lution of (?-aminothiaY,ol-~-y:L)acetic acid hydroch,Lor:ide
(0.~2 g) ancl dicycLohexy'Lcar'bodiim:ide~ (0.41 g) in dlraet'hyl-
`or~namide ( 20 m:l ) . 'llhe rn:ixture ;.s ~ tirred at roorn ternperature
and the resultirl~ pr~iplta-te i~ rernoved by suct:Lon filtxation.
The filtrate is mixed with ethylacetate (100 rnl) and chilled
water (100 ml), and the organic layer ls separated. The
organic layer is washed-with saturated aqueous sodium chloride
solution, dried over anhydrous sodium sulfate, and concentrated
~der reduced pressure to obtain white powder. Yield 0.51 g.
This product is coincided with the authentic sample obtained
above in ~MR and IR.
Example 6
Preparation of pivaloyloxymethyl 7~-[2-(2-amino-
thiazol~4-~rl)ace-tamido]-3-[[~1-(2-dimethylaminoethyl)-lH-
tetra~Ql-5-yl]thio]methyl]ceph-3-em-4-carboxylate dihydrochloride.
~ he above compound (IV) (0.57 g) is s-tirred with
sodium hydrogen carbonate (0.17 g) in the mixture of dichlor-
methane (15 ml) and water (15 ml). The organic layer is
separated, dried over anhydrous calcium chloride, and evaporated
under reduced pres3ure. The residue is dissolved in dichloro-
methane (15 ml), and chilled to -25C. To the solution is
added dropwise a solution of 4-chloro-3-oxo-butyrylchloride
(1.3 g) in dichloromethane (2.0 mi). After stirring for
20 minutes between -15 and -20C, to the mixture is added
aqueous sodium chloride solution. The organic layer is
separated, and washed with saturated aqueous sodium chloride
solution, and concentrated under reduced pressure. rrhe
residue is soli'dified by a mixture of ethyl acetate and
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~-` t~!f'~ . I'h~ L~ i V~l I 0,~ loxymc~thyl ~-(4~ Joro-~-oxo-
~:)utyry3,arrl:i.(1cl)-3~[ [ [1-(2-d irne-thy:la~rli.noe thy~l,)-:LH-tetrazol-5-y:l]thio]mcthyl~ceph-3-em-~-car~oxy:lci-te [forrnuIa (VI~], is
-taken up by suction fi:ltrat:ion. YLeld 0,2~ g.
IR (KBr, cm ]-): 17~0, :l748, ]6~0
To -the solution of the above compound (VI) (0.24 g)
in dichloromethane (20 m:L) is added a solution of thiourea
(0.38 g) in dimethylacetamide ~5 ml), and stirred for 3
hours at room temperature. To the mixtuYe is added water,
and -the aqueous layer is separated. It is adjusted to pH
6.0, and extracted with dichloromethane. On the organic
layer is layered water, adjusted to pH l.5 with 4N-HCl, and
the aqueous layer is separated. After removing dichloromethane
under reduced pressure, the aqueous layer is chromatographed
on Amberli~e XAD-II. The column is eluted successively with
O.OlN~HCl (120 ml~ and 5~ acetonitrile-O.OlN-HCl. The effluent
is lypophilized to give the objec-tive compound as white powder.
Yield 0.28 g. ~h1s product~is coincided with the authent1c
sample obtained a~ove ln NMR, IR and T~C.
.
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