Note: Descriptions are shown in the official language in which they were submitted.
~Z~8~
7566-720D
This is a divisional application of Serial No. 376,376 filed
April 28, 1981.
This invention relates to i.ntermediates useful for producing
novel Ampicillin esters, and to processes for the production of the
intermediates.
Ampicillin (aminobenzylpenicillin) obtained by acylating the
amino group of 6-aminopenicillanic acid (6-APA) with ~-aminophenylacetic
acid is a synthetic penicillin which is effective in oral administration.
Absorption of Ampicillin from the digestive tract, however, is not sufficient,
and this necessarily leads to administrati.on of large dosages for obtaining
the required concentration in blood, which in turn causes increased side-
effects.
To remove such a defect of Ampicillin, an attempt was made to
convert Ampicillin to an ester-type derivative thereby improving its
absorption from the intestinal tract. For example, Ampicillin pivaloyloxy-
methyl ester (Pivampicillin; see British Patent No.1,215,812~ and Ampicillin
phthalidyl ester (Talampicillin, see British Patent No. 1,36~,672) gives
comparable blood Ampicillin concentrations in oral administration to those
obtained by i.ntramuscular administration.
According to one aspect of the invention, there is provided
a process for preparing a 6-aminopenicillanic acid ester of the general
formula
(V)
o coofH - C= C - Rl
R2 () P
~( ~
-- 1 --
~23~1~
wherein Rl represents a hydrogen atom, a methyl group or an aryl group, and
R2 represents a hydrogen atom or may be taken together with Rl to form a
divalent carbon chain residue, or its acid addition salt~ which process
comprises:
(a) reacting 6-aminopenicillanic acid or its salt at the carboxyl
group with a compound of formula (III)
X-C~I-C - C-R
I I ! (III)
R2 ~
wherein Rl and R2 are as defined above and X is a halogen atom; or
(b) reacting 6-protected aminopenicillanic acid or its salt at the
carboxyl group with a compound of formula (III) as defined above and then
converting the protected amine group of the product to the free amine group; and
if required, converting the product to an acid addition salt thereof.
Another aspect of the invention provides the compound of formula (V)
as defined above.
In formula (V), Rl represents a hydrogen atom, a methyl group or
an aryl group. The aryl group is preferably an aromatic hydrocarbon group.
Preferred aromatic hydrocarbon groups are phenyl and substituted phenyl
groups, and the phenyl group is especially preferred. Examples of substituents
in the substituted phenyl groups are halogen, nitro, cyano and alkoxy. Thus, R
is preferably a hydrogen atom, a methyl group or a phenyl group.
R2 represents a hydrogen atom, or together with Rl, may form a carbon
chain residue. When R2 and Rl together form a divalent carbon chain residue,
Rl, R2 and the group C-C-CH to which Rl and R2 are bonded form a ring. The
divalent carbon chain residue is preferably an alkylene residue of 3 to 5 carbon
~2;~3~1
atoms so that the ring is 5- to 8-membered, particularly 6- or 8-membered.
Examples of preferred divalent carbon chain residues are -~CH2~3 and
2~5 '
The parent application concerns Ampicillin esters of the
general formula
r
O N- ~ COOCH-C _C-R
2 ~O~
wherein Rl and R2 are as defined above.
Specific examples of preferred Ampicillin esters of general
formula ~I) are
Ampicillin (5-methyl-2-oxo-1, 3-dioxolen-4-yl)-
methyl ester (Rl=methyl, R2=hydrogen),
- 2a -
~383~1
-3- 7566-720D
Ampicillin(2-oxo-1,3-dioxolen-4-yl)methyl ester
(Rl and R2= hydrogen),
Ampicillin(2-oxo-5-phenyl-1,3-dioxolen-4-yl)-
methyl ester (Rl=phenyl, R2=hydrogen),
Ampicillin(2,3-carbonyldioxy-2-cyclohe~en-1-
yl)ester (Rl and R2 together form the group
tCH2t3)' and
Ampicillin(2,3-carbonyldioxy-2-cycloocten-1-yl)-
ester (Rl and R2 together form the group tCH2t5)-
The acid addition salts of these Ampicillin
esters are, for example, salts of these esters with inorganic
acids such as hydrochloric acid, hydrobromic acid, hydriodic acid
and sulfuric acid, or salts of these with organic acids such as
citric acid and tartaric acid.
Investigations of the present inventors have shown that theAmpicillin esters or the acid addition salts thereof have very
desirakle properties as pharmaceuticals.
Specifically, in oral administration, the Ampicillin esters
are easily absorbed from the digestive tract, liberate Ampicillin
in vivo, and maintain a high Ampicillin concentration in blood
over long periods of time.
For example, thirty minutes after oral administration in
mice, Ampicillin (5-methyl-2-oxo-1,3-dioxolen-4-yl)methyl
ester hydrochloride and Ampicillin (2-oxo-5-phenyl-1,3-dioxolen
-4-yl)methyl ester hydrochloride show an Ampicillin concentration
in blood about 3 times as high as that attained by the
J
~;~38~
-4- 7566-720D
administration of Ampicillin and about 1.5 times as high as that
attained by the administration of Ampicillin phthalidyl ester,
and the high Ampicillin concentrations in blood are maintained
over a long period of time. (See Experiment 1 given hereinbelow.)
Such an excellent advantage of the Ampicillin esters of
this invention is believed to be due to the fact that while these
Ampicillin esters readily undergo enzymatic hydrolysis in vivo,
they have resistance to hydrolysis in gastric and intestinal
juices.
The rates of hydrolysis of Ampicillin (5-methyl-2-oxo-1,
3-dioxolen-4-yl)methyl ester hydrochloride and Ampicillin (2-oxo-
5-phenyl-1,3-dioxolen-4-yl)methyl ester hydrochloride in
simulated gastric and intestinal juices are about one half of
that of Ampicillin phthalidyl ester (see Experiment 2, (a) and
(b) hereinbelow).
Needless to say, this high chemical stability of the
penicillin esters of the invention is very beneficial not only in
bulk preparation and pharmaceutical preparation and also in actual
administration.
It is also noted that the above Ampicillin esters have low
toxicity (see Experiment 3 herein below).
Experiments 1 to 3 are described below for demonstrating
these advantages of these Ampicillin esters.
Experiment 1
Concentration in blood in oral administration (test Compounds)
A. Ampicillin (5-methyl-2-oxo-1,3-dioxolen-4-yl)methyl ester
hydrochloride.
33il 1
-5- 7566-720D
B. Ampicillin (2-oxo-5-phenyl-1,3-dioxolen-4-yl)methyl
ester hydrochloride.
C. Ampicillin phthalidyl ester hydrochloride (a known
compound used as a control; see British Patent No.
1,364,672)
D. Ampicillin trihydrate (control)
(Method of Experiment)
Each of the test compoinds was orally administered in a
dose of 50 mg/kg calculated as Ampicillin to four week old
mice (ddY, body weight about 20 g, five per group) which had
been caused to fast overnight (the amount is equivalent to 0.2 ml
of a Smg/ml aqueous solution of Ampicillin). The blood was taken
from the experimental animals periodically, and the concentration
of Ampicillin in the serum was measured by a bioassay method.
The blood Ampicillin level ratio was calculated from the following
equation.
Serum Ampicillin level in administration
of each of the compounds
Ampicillin A, B, C and D
level ratio Serum Ampicillin level in administration
of the compound D
(Result)
339~
-6- 7566-720D
Table 1
\ ~~~--_ Item Ampicillin level ratio
\ ~ , __ _~ _
Time of
Blood taking
(min.) 15 30~0 90 120 180
I Compound r i ~ ~
Compounds A 2.8 2.92.11.8 1.5 1.3
of formula
(I) _ B 2.4 2.82.4rl 9 _1.2 1.0
Known C 3.0 1.81.41.1 0.9 0.8
compound _
Control D 1.0 l.0l.0l.0 1.0 1.0
compound_ _
The results given in Table 1 clearly show that the compounds
of formula (I) show a high blood Ampicillin level over a longer
period of time than the known phthalidyl ester C.
Experiment 2
Hydrolyzability in acidic and basic media
(a) Hydrolyzability in an acidic medium (Test compounds)
Compounds A, B and C in Experiment l
(Method of Experiment)
Each of the test compounds was dissolved to a predetermined
concentration in an acidic aqueous medium (simulated gastric
juice) having a pH of 1.2 prepared by adding 2.0 g of sodium
chloride, 24 ml of lO~ hydrochloric acid and 3.2 g of pepsin to
lO00 ml of water. While the solution was shaken at 37C, it was
periodically sampled. The sampled solution was subjected to
3~
7--
high-speed liquid chromatography uslng a reversed phase partition
column, and the hydrolysis ratio of the compound was determined
from a decrease in the peak height of the compound in the
chromatogram.
(Results)
Table 2
Item Hydrolysis ratio (%)
Time of _ _
sampling (hrs) 1 2 4 6 20
compound
_ 7 15 20 28 52
Compounds
of formula B 9 20 30 35 65
Known C 18 31 43 55 100
compound
(b) Hydrolyzability in a basic medium (Test compounds)
Compounds A, B and C of Experiment 1
(Method of Experiment)
The pxocedure of (a) was repeated except that a basic
aqueous medium (simulated intestinal juice) having a pH of 7.50
prepared by adding 35.8 g of disodium phosphate. 6.0 ml of 10%
hydrochloric acid and 2.8 g of pancreatin to 1000 ml of water was
used instead of the acidic aqueous medium.
(Results)
:~2383~
-7a 7566-720D
Table 3
_
Item Hyd rolysi s rati ) (%)
Time of
sampling 5 10 20 30 60
Test (min.)
compound
Compounds A 16 32 48 65 80
of formula
(I) B 18 37 53 70 90
_
Known C 39 61 90 95 100
com~ound
_ l
The results given in Tables 2 and 3 demonstrate that the
compounds of formula (I~ have higher chemical stability in acidic
and basic conditions than the known phthalidyl ester C.
Experiment 3
Acute toxicity:-
The acute toxicity values (LD50) of the compounds A and Bin experiment 1 administered as an aqueous solution were measured
using the same ddY mice as used in Experiment 1. The results
are shown in Table 4. The results show that the compounds of
formula (I) have low toxicity.
Table 4
_ . .
Item LDsn (mg/k~
Route
Test oral Intraperitoneal Intravenous
Compound
A >5,000 1,430 557
B >5,000 _1,768 270
123~31~
-7b- 7566-720D
Prodrugs such as Ampicillin pivaloyloxymethyl ester
or Ampicillin phthalidyl ester have been known as
~2319,3~L~
orally administrable Ampicillin. The ester group of the Ampicillin ester of
formula (I) (i.e., (2-oxo-1,3-dioxolen-4-yl) methyl group) is shown by a
formula below in comparison with those of the known prodrugs.
Ester group
o
Ampicillin puvaloyloxy -C~l20-C-C(CH
methyl ester
Ampicillin phthalidyl b t
ester
Ampicillin ester -Ci-l-C=C-R
of formula ~ f ¦
R2
It is clear therefore that the ester group of the Ampicillin ester
of formula (I) quite differs from those of the known Ampicillin esters. It
is surprising that the Ampicillin esters of formula (I) have the aforesaid
excellent properties as pharmaceuticals over these known Ampicillin esters.
The compounds of general formula III are novel and are the
subject of Canadian Patent Application Serial No.376,391. In formula III,
Rl and R2 are as defined above with regard to formula V, and X represents a
halogen, preferably chlorine, bromine and iodine.
Examples of the compound of formula III are 4-chloromethyl-1,3-
dioxolen-2-one, 4-bromomethyl-1, 3-dioxolen-2-one, 4-chloromethyl-5-phenyl-1,3-
dioxolen-2-one, 4-bromomethyl-5-phenyl-1~3-dioxolen-2-one, 4-chloromethyl-5-
methyl-1,3-dioxolen-2-one, 4-bromomethyl-5-methyl-1,3-dioxolen-2-one, 4-iodo-
methyl-5-methyl-1, 3-dioxolen-2-one, 3-chloro-1,2-carbonyldioxycyclohexene, 3-
bromo-l, 2-carbonyldioxycyclohe~ene, 3-chloro-1,2-carbonyldioxycylooctene, and
3-bromo-1, 2-carbonyldioxycyclooctene.
-- 8 --
~Z3~3~
These compounds can be produced by reacting a compound of the
formula
CH2-C.~ C-
~R2
o
wherein Rl and R2 are as defined above, with halogenating agents, for example
allylic halogenating agents such as chlorine, bromine, N-bromosuccinimide and
N-chlorosuccinimide. The compounds of general formula III and their production
are described in the specification of a patent application filed by the same
applicants as the present application on the same date.
The 6~aminopenicillanic acid ester of general formula (V),
which is the subject matter of the present application, is useful for producing
the Ampicillin ester of general formula (I) mentioned before. The compound
of formula (V) or its acid addition salt may be reacted with a carboxylic
acid of the general formula
CHCOOH (VI)
-- A
wherein A is a protected amino group or a group convertible to an amino group,
or its reactive derivative at the carboxyl group; thereafter, if required,
when the resulting compound has the protected amino group or the group
convertible to an amino group, eliminating the protective group from the
protected amino group, or converting said convertible group to an amino group,
and if further required, converting the product to an acid addition salt thereof.
~Z3~,3~1
Preferred examples of the compound of general formula (V) are:
(2-oxo-1,3-dioxolen-4-yl)methyl 6-aminopenicillanate,
(5-methyl-2-oxo-1, 3-dioxolen-4-yl)methyl 6-amino-
penicillanate,
(2-oxo-5-phenyl-1, 3-dioxolen-4-yl)methyl 6-amino-
penicillanate,
(2,3-carbonyldioxy-2-cyclohexen-1-yl) 6-amino-
penicillanate,
(2,3-carbonyldioxy-2-cycloocten-1-yl)6-amino-
penicillanate,
and
acid addition salts of these esters.
The compound of general formula (V) can be produced by reacting
6-aminopenicillallic acid or its salt or its reactive derivative at the
carboxyl group with the compound of general formula (III); or by reacting
6-protected aminopenicillanic acid or its salt or its reactive derivative at
the carboxyl group with the compound of general formula (III) and then
converting the protected amino group of the reaction product to the amino
group.
The former reaction can be performed preferably by reacting 6-
aminopenicillanic acid or its salt at the carboxyl group with an equimolar
amount, or a molar excess, of the compound of general formula (III) in an
inert organic solvent such as tetrahydrofuran, dioxane or acetone in the
optional presence of a base (when 6-aminopenicillanic acid is used, the
presence of a base is preferred) at a temperature of from about 0C to room
temperature.
- 10 -
1~3~33~:~
The latter reaction can be performed preferably by reacting 6-
protected aminopenicillanic acid such as 6-aminopenicillanic acid having the
amino group at the 6-position protected with an acyl group or trityl group,
or 6-aminopenicillanic acid having the amino group at the 6-position protected
as a Schiff base, or its salt at the carboxyl group, for example 6-phenyl-
acetylaminopenicillanic acid (benzylpenicillin), with the compound of
general formula (III) under the same conditions as in the first-mentioned
process, thereafter reacting the resulting 6-protected aminopenicillanic acid
ester with phosphorus pentachloride and a lower alcohol such as methanol at
the temperature of dry ice-acetone in the presence of a basic compound such as
N-methylmorpholin, quinoline and triethylamine, and thereafter causing water
to act on the resulting imino ether to hydroly~e it.
For producing the Ampicillin esters of formula ~I), the compound
of general formula ~V) or its acid addition salt is first reacted with the
carboxylic acid of general formula ~VI) or its reactive derivative.
The acid addition salt of the compound of general formula ~V)
may be a mineral acid salt or an organic acid salt, for example such a
mineral saltasahydrochloride or hydrobromide, or such an organic acid salt
as a para-toluenesulfonate.
Acid halides, acid anhydrides and mixed acid anhydrides are
preferably used as the reactive derivative of the carboxylic acid of
general formula ~VI).
~23~3~1
-12- 7566-720D
The reaction of the compound of general formula (V) or its
acid addition salt with th- carboxylic acid of general formula
(VI) is carried out in the presence of a dehydrocondensing agent
such as dicyclohexyl carbodiimide (DCC) or a mixture of DCC and
l-hydroxybenzotriazole, preferably in a solvent consisting
substantially of an aprotic inert organic solvent such as dimethyl
formamide, dimethyl sulfoxide, methylene chloride, dioxane and
tetrahydrofuran at a temperature of not more than 50C.
The reaction of the compound of general foxmula (V) or
its acid addition salt with the reactive derivative of the
carboxylic acid of general formula (VI) is carried out preferably
in a solvent consisting substantially of an aprotic inert organic
solvent such as dimethyl formamide, dimethyl sulfoxide, methylene
chloride, dioxane tetrahydrofuran and acetone at a temperature
of not more than 50C, When an acid addition salt of the compound
expressed by general formula (V) is used, the reaction is
preferably carried out in the presence of a base such as
triethylamine.
The reactive derivative of the carboxylic acid of general
formual (VI) used in the above reaction is preferably an acid
halide such as an acid chloride when the group A in general
formula (VI) is a protected amino group in the form of a salt
with a mineral acid. The acid halide of the compound of general
formula (VI) having such a group A can be conveniently produced
by treating the compound of general formula (VI) having such a
group A with a halogenating agent such as thionyl chloride,
~3~
-12a- 7566-720D
phosgene or phosphorus pentachloride because such group A is
stable to acids.
The reactive derivative of the compound of general
formula (VI) is which the group A is a protected amino group in
the ~orm of a Schiff base or an enamine group is preferably an
acid anhydride or mixed acid anhydride. This reactive derivative
can be produced conveniently by treating a salt, such as
a trialkylamine salt,
{'~ ~
~23~3~
of the carbo~ylic acid of general formula (VI) in which the group A is such
a protected amino group, with, for example, an alkyl haloformate such as
ethyl chloroformate and isobutyl chloroformate.
The reaction between the compound of general formula (V) or
its acid addition salt and the carboxylic acid of general formula (VI) or
its reactive derivative gives the compound of general formula (IV),
> A~ N j\ C113 (IV)
o ` C00-CH-C_ - C-R
R~0\ ~,0
wherein Rl and R2 are as defined above and A' represents a protected amino
group~ a group convertible to an amino group, or an amino group. When the
compound of formula (IV) has as the radical A' a protected amino group or
a group convertible to an amino group, the protecting group is removed from
the protected amino group, or the convertible group is converted to an amino
group and if desired, the product is converted to its acid addition salt.
Thus, the Ampicillin ester of general formula (I) or its acid addition salt
is formed.
According to a preferred process for producing the Ampicillin ester
of general formula (I) or its acid addition salt, a compound of general formula
(VI) in which A is a Schiff base group or an enamine group is reacted with the
compound of general formula (V), thereafter the Schiff base group or the enamine
group (A) of the resulting compound is converted to an amino group. Preferably
for producing an acid addition salt, such as hydrochloride, of the Ampicillin
ester of general formula (I), a compound of~general formula (VI) in which A is in
the form of an acid addition salt such as a hydrochloride is reacted with the
compound of general formula (V).
- 13 -
~23~3~
After the reaction, the Ampicillin of general formula (:[) or its
acid addition salt can be isolated and purified in a customary manner.
The Ampicillin ester of general formula (I) or its pharmaceutically
acceptable acid addition salt is converted back to Ampicillin in ivo when
administered to an animal.
The Ampicillin ester of general formula (I) or its pharmaceutically
acceptable acid addition salt may be used as an active ingredient of anti-
bacterial agents.
The antibacterial agent may consist only of the Ampicillin ester of
general fo-rmula (I) or its pharmaceutically acceptable acid addition salt, or
may contain a pharmaceutically acceptable carrier.
The pharmaceutically acceptable carrier may be those carriers which
can be used in formulating Ampicillin. Examples are starch, lactose,
hydroxypropyl,celluloSe,crystalline cellulose, magnesium stearate, and calcium
stearate.
The antibacterial agent may be administered orally, for example. It
may be in a unit dosage form for oral administration, such as tablet (sugar-
coated tablets), capsules, granules and powder.
The antibacterial agent may be administered to man and other animals
in a dose of 1 to 50 mg/kg body weight/day calculated as the Ampicillin ester
(I) or its pharmaceutically acceptable salt.
l'he following Examples illustrate the present invention, as well as
that of the parent application.
Exa le 1
mp
(1) Production of 4-bromomethyl-5-phenyl-1, 3-dioxolen-2-one:-
In 150 ml of carbon tetrachloride was dissolved 2.4 g of 4-methyl-5-
phenyl-l, 3-dioxolen-2-one(synthesized by the method described in Liebichs
- 14 -
~23~3~
Annalen der Chemie, Vol.764, pages 116-124, 1972). N-bromosuccinimide (2.9 g)
and a catalytic amount of ~, ~'-azobisisobutyronitrile were added to the
solution, and the mixture was heated under reflux for 90 minutes. The
reaction mixture was concentrated to one half of its volumc, ~md the insoluble
material was separated by filtration. The filtrate was concentrated,
and the residue was recrystallized from
- 14a -
~23~3~
-15- 7566-720D
a mixture of benzene and cyclohexane to give 2.3 g (yield 66%) of
colorless needles having a melting point of 90.5 to 91.5C.
This product had the following properties.
Elementalanalysis, molecular formula ClOH7BrO3:
Calculated (%): C, 47.09; H, 2.77; Br, 31.33
Found (%): C, 47.22; ~, 2.64; Br, 31.29
IR (KBr): near 1825 cm 1 (v
NMR (CC14,~(ppm)):
4.35 (-CII2Br, s), 7.40 (benzene ring, s)O
From these data, the product was identified as the
title compound.
(2) Production of Ampicillin (2-oxo-5-phenyl-1,3-dioxolen-4-yl)
methyl ester hydrochloride:-
Ampicillin trihydrate (500 mg) was dispersed in 6 ml
of dimethyl formamide, and 125 mg of potassium hydrogen carbonate
was added. The mixture was cooled to 0C and stirred. Benzalde-
hyde (0.25 ml) was added, and the mixture was stirred at 0C for
2.5 hours. Then, 125 mg of potassium hydrogen carbonate and
320 mg of 4-bromomethyl-5-phenyl-1,3-dioxolen-2-one were added,
and the mixture was further stirred at 0C for 3 hours.
After the reaction, the reaction mixture was poured into
the ice water and extracted with ethyl acetate. The extract was
washed three times with ice water. The ethyl acetate layer was
concentrated under reduced pressure to form a syrup. The syrup
was dissolved in 4 ml of acetonitrile, and the pH of the
solution was adjusted to 2.0 with dilute hydrochloric acid. The
383~1
-15a- 7566-720D
solution was then stirred at 0C for 30 minutes.
~ ater (10 ml) was added, and the mixture was concentrated
under reduced pressure to distill off acetonitrile. The aqueous
layer was repeatedly washed with ethyl acetate, and saturated
with sodium chloride. The separated oily substance was
extracted with 50 ml of methylene chloride, washed with a
saturated aqueous solution of sodium chloride and dried over
anhydrous sodium sulfate. The dried organic layer was
concentrated until the amount
~L23~131~
of mcthylene chloride decreased to one half. Isopropyl alcohol (30 ml) was
added, and the mixture was again concentrated under reduced pressure to give a
colorless solid. The solid was collected by filtration, and washed successively
with isopropyl alcohol and ether to give 320 mg (yield 46.4%) of Ampicillin
(2-oxo-5-phenyl-1,3-dioxolen-4-yl)methyl ester hydrochloride as a colorless
amorphous solid.
Melting point: 140C (decomp.)
Elemental analysis, molecular formula
C26H25N307S HCl 2H2
Calculated ~%): C, 52.39; H, 5.07; N, 7.05; S, 5.38;
Found %: C, 52.17; H, 4.83; N, 7.31; S, 5.64.
IR (KBr): 1830 cm 1 (cyclic carbonate), 1785 cm 1 (~-lactam),
1760 cm 1 (ester), 1690 cm 1 (amide).
NMR (DMSO-d6, ~(ppm)): 1.32 and 1.45 (6H, methyl at the 2-position,
s), 4.44 (lH, proton at the 3-position, s), 5.12 ~lH, benzyl proton! s), 5.31
(2H, -CH2-1C=IC-C6H5, s), 5.4-5.6 (2H, pTotons at the 5- and 6-positions, m),
0~0
7.3-7.6 (lOH, protons on the benzene ring, m), 8.8 (3H, -NH3), 9.3 (lH, -CONH-,
d)-
The resulting Ampicillin ester hydrochloride was incubated in 40%
mouse blood in pH 7.4 phosphate buffer at 37C for 10 minutes, and then sub-
jected to bioautography. It was found to be completely converted to Ampicillin.
Example 2
(1) Production of 4-bromomethyl-5-methyl-1,3-dioxolen-2-one:-
- 16 -
;3~1
-17- 7566-720D
In 150 ml of carbon tetrachloride was dissolved 3.42 g
of 4,5-dimethyl-1,3-dioxolen-2-one (synthesized by the method
of described in Te-trahedron Letters, 1972, pages 1701-1704).
N-bromosuccinimide (5.34 g) and a catalytic amount of ~
azobisisobutyronitrile were added to the solution, and the
mixture was heated under reflux for 15 minutes. The reaction
mixture was concentrated to one half of its volume, and the
insoluble material was removed by filtration. The filtrate was
concentrated, and the syrupy residue was distilled under reduced
pressure to give 4.2 g (yield 73%) of a colorless liquid having
a boiling point of 115-120C/5 mm. The product had the following
properties.
Elemental analysis, molecular formula C5H5BrO3:
Calculated (~): C, 31.12; H, 2.61; Br, 41.40
Found (%): C, 31.30; H, 2.49; Br, 41.31
IR (neat): near 1825 cm 1 (vC=O)
NMR (Ccl4r~(ppm)):
2.10 (-CH3, s), 4.10 (-CH2Br, s).
From these data, the product was identified as the title
compound.
(2) Production of Ampicillin (5-methyl-2-oxo-1,3-dioxolen-4-yl)
methyl ester hydrochloride:-
Ampicillin trihydrate (500 mg) was dispursed in 6 ml ofdimethyl formamide, and 125 mg of potassium bicarbonate was
added. The mixture was cooled to 0C, and 0.25 ml of
benzaldehyde was added. The mixture was stirred at 0C for 2.5
~2383~1
-17a- 7566-720D
hours. Then, 125 mg of potassium bicarbonate and 250 mg of
4-bromomethyl-5-methyl-1,3-dioxolen-2-one were added, and the
mixture was stirred at 0C for 3 hours. After the reaction, the
react.ion mixture was poured into ice water, The precipitated
solid was extracted with 30 ml of ethyl acetate. The organic
layer was washed with 20 ml of water three times, and dried over
anhydrous magnesium sulfate. The ethyl acetate was distilled
off under reduced pressure to give a yellow syrup.
~23~33~
The resulting syrupy residue was dissolved in 4 ml of acetonitrile
and the solution was adjusted to pli 2.0 with dilute hydrochloric acid. The sol-
ution was then stirred at 0C for 30 minutes. Water (10 ml) was added, and t'ne
acetonitrile was distilled off under reduced pressure. The aqueous layer was
washed repeatedly with ethyl acetate, and then saturated with sodium chloride.
The separated oily substance was extracted with 50 ml of methylene chloride,
and ~ashed with a saturated aqueous solution of sodium chloride. The methylene
chloride layer was dried over anhydrous sodium sulfate, filtered and concentrat
ed under reduced pressure to one half of its volume. To the solution isopropyl
alcohol (30 ml) was added, and the mixture was again concentrated under reduced
pressure to give a colorless amorphous solid.
The solid was collected by filtration and washed with isopropyl
alcohol and ether to give 312 mg (yield 50.6%) of Ampicillin (5-methyl-2-oxo-
1,3-dioxolen-4-yl)methyl ester hydrochloride as a colorless amorphous solid.
The product had the following properties.
Appearance: Colorless amorphous solid
Melting point: 145C (decomp.)
~lemental analysis, molecular formula
C21H23N307SrHCl H2
Calculated (%): C, 48.88; H, 5.08; N, 8.14; S, 6.21;
Found (%): C, 48.51; H, 5.15; N, 8.02; S, 6.44;
IR (KBr): 1825 cm (cyclic carbonate), 1785 cm 1 (~-lactam),
1750 cm l (ester), 1690 cm 1 (amide).
NMR (D~IS0-d6, ~(ppm)): 1.33 and 1.48 (6H, methyl at the 2-position,
s), 2.19 (311, Cl-l3-1C=C-, s), 4.43 (lH, proton at the 3-position, s), 5.11
o
- 18 -
~ 23~3~
(2H, CH3-C=C-CH2-, s), 5.16 (1~1~ benzyl proton, 5), 5.43-5.65 (21-1, protons at o~f
o
the 5- and 6-positions, m), 7.3-7.6 (Sl-l, protons on the benzene ring, m). 8.95~3H, -NH3), 9.4 (lH, CON~I-, d).
The resultir.g Ampicillin ester hydrochloride was incubated in 40%
mouse blood in p~l 7.4 phosphate buffer at 37C for 10 minutes, and then subject-
ed to bioautography. It was found to be completely converted to Ampicillin.
Example 3
~1) Production of 3-bromo-1,2-carbonyldioxycyclohexene:
In 80 ml of carbon tetrachloride was dissolved 2.15 g of 1,2-
carbonyldioxycyclohexene (synthesized by the method described in Tetrahedron
- Letters, 1972, pages 1701 1704). N-bromosuccinimide ~2.3 g) and a catalytic
amount of ~,~'-azobisisobutyronitrile were added to the solution, and the
mixture was heated under reflux for 20 minutes. The reaction mixture was
cooled, and filtered. The filtrate was concentrated at a low temperature to
give 3.2 g of a pale brown liquid as a residue. The product showed the follow-
ing properties.
IR (neat): near 1825 cm (v O)
NMR (CDC13, ~(ppm)): 5.0 (=C-CH-Br, m), l.3-3.0 (cyclic proton, m).
From these data, the product was identified as the title compound.
(2) Production of Ampicillin (2,3-carbonyldioxy-2-cyclohexen-1-yl)
ester hydrochloride:
By the same method as shown in Example 1, (2), 256 mg of Ampicillin
(2,3-carbonyldioxy-2-cyclohexen-1-yl) ester h~ydrochloride was obtained as a color
less amorphous solid from 2 g of Ampicillin trihydrate and 1 g of 3-bromo-1,2-
carbonyldioxycyclohexene (yield 10.2%).
Appearance: colorless amorphous solid
- lg -
12383~
~lelting point: 140C (decomp.j
IR (KBr): 1830 cm (cyclic carbonate), 1780 cm (~-lactam), 1750
cm (ester), 1690 cm (amide).
The resulting Ampicillin ester hydrochloride
- 20 -
123~
-21- 7566-720D
was incubated in 40% mouse blood in pH 7.4 phosphate buffer at
37C for 10 minutes, and then subjected to bioautography. It
was found to be completely converted to Ampicillin.
Example 4
(1) Production of 4-bromomethyl-1,3-dioxolen-2-one:-
In 200 ml of carbon tetrachloride was dissolved 8.6 gof 4-methyl-1,3-dioxolen-2 one (synthesized by the method
described in U.S. Patent No. 3,020,290). N-bromosuccinimide
(17.8 g) and a catalytic amount of ~,~'-azobisisobutyronitrile
were added to the solution, and the mixture was heated under relux
for 90 minutes. The reaction mixture was worked up in the same
way as in Example 2 to give 5.2 g (yield 33.6%) of a colorless
liquid having a boiling point of 94C/3 mm. The product had
the following properties.
Elemental analysis, molecular formula C4H3BrO3:
Calculated (%): C, 26.84; H, 1.69; Br, 44.65
Found (%): C, 26.94; H, 1.66; Br, 44.60
IR (neat): near 1830 cm 1 (v
NMR (CC14,~(ppm)):
4.10 (-CH2Br, s), 7.00 (=CH-O-, s).
From these data, the product was identified as the
title compound.
(2) Production of Ampicillin (2-oxo-1,3-dixolen-4-yl)-methyl
ester hydrochloride:-
Ampicillin trihydrate (2 g) was dispersed in 24 ml ofdimethyl formamide, and 500 mg of potassium hydrogen carbonate
~23~3~
-22- 7566-720D
was added. The mixture was cooled to 0C, and 1 ml of
benzaldehyde was added. The mixture was stirred to 0 to 5C for
3 hours. To the mixture were added 500 mg of potassium hydrogen
carbonate and 1 g of 4-bromomethyl-1,3-dioxolen-2-one, and the
mixture was stirred at 0 to 5C for 6 hours.
After thereaction, the reaction mixture was poured into
ice water, and extracted with ethyl acetate. The extract was
washed with ice water, and the ethyl acetate layer was
concentrated under reduced pressure to remove ethyl acetate. The
resulting syrup was dissolved in 10 ml of acetonitrile. The
solution was adjusted to pH 2.0 with dilute hydrochloric acid, and
stirred at 0C for 20 minutes.
Water (20 ml) was added, and the mixture was concentrated
under reduced pressure to remove acetonitrile. The aqueous layer
was repeatedly washed with ethyl acetate, and saturated with
sodium chloride to precipitate and orange gum-like substance.
The a~ueous layer was removed by decantation. The gum-like
substance was dissolved in methanol, decolorized with activated
carbon, cooled to 0C, and poured into vigorously stirred ether to
precipitate a pale orange solid. The solid was collected by
filtration, and washed with a mixture of ether and methanol to
give 600 mg (yield 26%) of Ampicillin (2-oxo-1,3-dioxolen-4-yl)
methyl ester hydrochloride as a pale orange amorphous solid.
Melting point: 130C (decomp.)
IR (KBr): 1835 cm 1 (cyclic carbonate), 1790 cm 1
~2383~
-22a- 7566-720D
(~-lactam), 1750 cm 1 (ester), 1690 cm 1
(amide)
NMR (deuterium oxide,~ (ppm)):
1.36 (6H, methyl at the 2-position, s), A.58
(lH, proton at the 3-position, s), 5.11 (2H,
-CH2-C=CH, s), 5.23 (lH, benzyl proton, s),
0\ ~0
5.49 (lH, proton at the 5-position, d, J=2.0 Hz),
5.58 (lH, proton at the 6-position, d, J=2.0 ~Iz)
7.5 (6H, -C=CH and benzene proton, m),
o~o/o
The resulting Ampicillin ester hydrochloride
~23~3~
was incubated in 40% mouse blood in pl-l 7.4 phosphate buffer at 37C for 10
minutes, and then subjected to bioautography. It was found to be completely
converted to Ampicillin.
Exampl
(1) Production of Benzylpenicillin (5-methyl-2-oxo-1,3-dioxolen-4-
yl)methyl ester:
Ten grams of benzylpenicillin potassium salt was dispersed in
50 ml of dimethyl formamide, and with ice cooling, 5Z0 mg of potassium hydrogen
carbonate and 5.2 g of 4-bromomethyl-5-methyl-1,3-dioxolen-2-one were added,
then the mixture was stirred at oC for 4 hours. The reaction mixture was
poured into ice water, and the precipitated solid was collected by filtration.
It was dissolved in ethyl acetate, washed with a dilute aqueous solution of
sodium hydrogen carbonate and then repeatedly with ice water. The ethyl acetate
layer was then dried over anhydrous magnesium sulfate, and the solvent was
distilled off under reduced pressure. There was obtained 12.5 g (yield 94%) of
benzylpencillin (5-methyl-2-oxo-1,3-dioxolen-4-yl)methyl ester as pale yellow
amorphous solid.
IR (KBr): 1825 cm (cyclic carbonate), 1785 cm (~-lactam), 1750
cm 1 (ester), 1670 cm 1 (amide)
NMR (CDC13, ~(ppm)): 1.37 and 1.42 (611, methyl at the 2-position, s),
2.13 (3H, -IC=C-CH3, s), 3.72 (2H, -CH2-C6H5, s~, 4.29 (li-l, proton at the 3-
O~f
o
position, s), 4.80 (2H, -CH2-1C=IC-, s), 5.3-5.6 (2H, protons at the 5- and 6-
O O
O
positions, m), 5.59 (11l, proton at the 5-position, d), 6.16 (lH, NH, d), 7.14
(5H, protons on the benzene ring, s).
~239,3~1
~2) Production of (5-metllyl-2-oxo-1,3-dioxolen-4-yl)methyl 6-amino-
penicillanate hydrochloride:
Phosphorus pentachloride (5.9 g) was dissolved in dry methylene
chloride (30 ml), and 6.3 ml of quinoline was added. The solution was cooled
to -30C witn dry ice-acetone. With vigorous stirring, 11 g of the above
benzylpenicillin (2-oxo-5-methyl-1,3-dioxolen-4-yl)methyl ester dissolved in drymethylene chloride (10 ml) was added dropwise, and the mixture was stirred at
this temperature for 35 minutes. Propyl alcohol (20 ml) was added dropwise
over 5 minutes, and the mixture was stirred for 30 minutes. With vigorous
stirring, 20 ml of a saturated solution of sodium chloride was added dropwise
and the mixture was stirred for about an hour. Then, the methylene chloride
layer was separated, washed with saturated sodium chloride solution, dried over
anhydrous sodium sulfate and evaporated to dryness to afford a gum. The gum was
~ washed with n-hexane and then with ethyl acetate to give 6.6 g (yield 72%) of
(5-methyl-2-oxo-1,3-dioxolen-4-yl)methyl 6-amino-penicillanate hydrochloride
as a pale yellow amorphous substance.
~lelting point: 115-120C (decomp.)
Elemental analysis, molecular formula
cl3~ll6N26S H51 1/2H2
Calculated (%): C, 41.77; H, 4.S5; N, 7.49;
Found (%): C, 41.91; H, 4.77; N, 7.67.
IR (KBr): 1825 cm (cyclic carbonate), 1790 cm 1 (~-lactam),
1750 cm 1 (ester).
N~IR (D~iS0-d6, ~(ppm)): 1.45 and 1.66 (6H, methyl at the 2-position,
s), 2-22 (3H, -IC=lC-Ci-l3, s), 4.57 (lH, proton at the 3-position, s), 5.0-5.2 (3H,
O O
- 24 -
~L~3~,3~1
proton at the 6-position, -C1l2-C=C-), 5.56 (li~, proton at the 5-position, d).
O~ O
o
(3) Production of D-(-)-phenylglycyl chloride hydrochloride:
Separately, 10 g of D-(-)-phenylglycine was added to 250 ml of
methylene chloride. The mixture was cooled to 0C, and by passing hydrogen
chloride gas, the hydrochloride of the phenylglycine was formed. Phosphorus
pentachloride (20 g) was added, and the mixture was stirred at 0 to 5C for 4
hours. The solid precipitated was collected by filtration, and repeatedly
washed with methylene chloride to give 13.1 g ~yield 90%) of D- (-) -phenylglycyl
chloride hydrochloride as a colorless amorphous solid.
~4) Production of Ampicillin (5-methyl-2-oxo-1,3-dioxolen-4-yl)-
methyl ester hydrochloride:
~ Two-hundred milligrams of the 6-aminopenicillanic acid ester hydro-
chloride obtained as above was dispersed in 10 ml of methylene chloride, and
50 mg of potassium hydrogen carbonate was added. The mixture was stirred at
0C for 15 minutes. Then, 110 mg of the acid chloride obtained as above was
added, and the mixture was stirred for 2 hours and then for another 2 hours at
room temperature.
After the reaction, the solid was separated by filtration, and the
filtrate was concentrated under reduced pressure. The resulting syrup was
dissolved in water, and washed with ethyl acetate. The aqueous layer was
saturated with sodium chloride, and the separated oily substance was extracted
with methylene chloride. The extract was washed with a saturated aqueous
solution of sodium cl~loride and concentrated until the amount of methylene
chloride decreased to half. Upon addition of isopropyl alcohol, a colorless
solid was precipitated. The solid was collected by filtration and washed with
isopropyl alcohol and etl-er to give 132 mg (yield 54~) of Ampicillin (5-methyl-
- 25 -
~L23~3~1
2-oxo-1,3-dioxolen-~-yl)methyl ester hydrochloride as an amorphous solid.
The melting point and spectroscopic data of this product were identi-
cal with those of tl-e product obtained in Example 2 (2).
Example 6
Production of Ampicillin ~2-oxo-5-phenyl-1,3-dioxolen-4-yl)methyl
ester hydrochloride:
By the samc method as shown in Example 5, (2-oxo-5-phenyl-1,3-dioxolen-
4-yl)methyl 6-aminopenicillanate hydrochloride was obtained in a yield of 74%
from 4-bromomethyl-5-phenyl-1,3-dioxolen-2-one and benzylpenicillin potassium
salt.
Melting point: 105-110C (decomp.)
Elemental analysis, molecular formula
C18H18N206S Hcl`l/2ll2
~ Calculated (%): C, 49.60; H, 4.62; N, 6.42;
Found (%): C, 49.5S; H, 4.56; N, 6.65.
IR (KBr): 1830 cm (cyclic carbonate), 1790 cm (~-lactam), 1755
cm 1 (ester).
NMR (DMS0-d6, ~(ppm)): 1.46 and 1.63 (6H, methyl at the 2-position,
s), 4.55 (lH, proton at the 3-position, s), 4.95 (lH, proton at the 6-position,
d)J 5.36 (2H, -CH2-iC=IC-, s), 5.53 (lH, proton at the 5-position, s), 7.4-7.75
O~ O
o
(5H, aromatic protons).
From 200 mg of the resulting ester hydrochloride and 95 mg of D-(-)-
phenylglycyl chloride hydrochloride, 148 mg (yield 56%) of Ampicillin (2-oxo-
5-phenyl-1,3-dioxolen-4-yl)methyl ester hydrochloride was obtained as a color-
less amorpllous solid.
The melting point and syectroscopic data of this product were identi-
38~
cal with those of the product obtained in Example 1 (2).
Example 7
(1) Production of (5-methyl-2-oxo-1,3-dioxolen-4-yl)methyl G-amino-
penicillanate p-toluenesulfonate:
In 100 ml of dimethyl formamide was dissolved 13 g of 6~-tritylamino-
penicillanic acid synthesized by the method described in J. Am. Chem. Soc. 84,
2983 (1963). The solution was cooled to 0 to 5C, and 3 g of potassium hydro-
gen carbonate and 6 g of 4-bromomethyl-5-methyl-1,3-dioxolen-2-one were added.
The mixture was stirred at the above temperature for 3 hours. After the reac-
tion, the reaction mixture was poured into ice water. The precipi~ated yellow
solid was extracted with 300 ml of ethyl acetate. ~ne ethyl acetate layer was
washed several times with a saturated aqueous solution of sodium chloride,
~ dried over anhydrous magnesium sulfate and concentrated under reduced pressure
to give a yellow syrup. The syrup was dissolved in 80 ml of ethyl acetate, and
with ice cooling, 5.2 g of p-toluenesulfonic acid was added. The mixture was
stirred under ice cooling for 1 hour, whereupon a colorless solid precipitated.
The solid was collected by filtration and well washed with ethyl acetate to
give 8.3 g (yield 60%) of the title compound.
Melting point: 130-138C (decomp.)
Elemental analysis, molecular formula
, Cl3lll6N2o6s~cH3c6H4so3H
Calculated (%): C, 47.99; H, 4.83; N, 5.60
Found (%): C. 47.31; H, 4.82; N, 6.00.
IR (KBr): 1820 cm (cyclic carbonate), 1780 cm (3-lactam), 1760
cm 1 ~ester)
N~IR (DMS0-d6, ~(ppm)): 1.40 and 1.59 (611, methyl at the 2-position,
s), 2.12 (3H, -C=CI-CH3, s), 4.46 (lH, proton at tlle 3-position, s), 4.90-5.10
O O
- 26a -
~23~3~
(3~1, proton at the G-posit:ion and -Cl12-l=C , m), 5.41 (1~1, proton at the 5-
O O
position, d), 2.24 (311, C~13 ~ -S03~, s), 6.97 and 7.38 (4H, aromatic
- 26~ -
~23~33~
-27- 7566-720D
protons C~3 ~ SO3~ , d).
(2) Production of Amplcillin (5-methyl-2-oxo-1,3-dioxolen-4-yl)
methyl ester hydrochloride:-
Five grams of (5-methyl-2-oxo-1,3-dioxolen-4-yl)methyl
6-aminopenicillanate p-toluenesulfonate was suspended in 300ml
of ethyl acetate. To the suspension was added at 0C 200 ml of
a 2% aqueous solution of sodium hydrogen carbonate cooled with
ice. The mixture was vigorously stirred. The ethyl ace~ate
layer was separated, washed with ice water, dried at 0C over
anhydrous magnesium sulfate, and concentrated under reduced
pressure to give a pale yellow syrup. The syrup was dissolved
in 50 ml of methylene chloride. The solution was cooled to
0C, and 1 g of potassium hydrogen carbonate and 2.1 g of D-(-)-
phenylglycyl chloride hydrochloride were added, and the mixture
was stirred at 0C for 4 hours. After the reaction, the
insoluble material was separated by filtration, and the filtrate
was concentrated under reduced pressure. The resulting syrup
was dissolved in water and washed with ethyl acetate. The
aqueous layer was saturated with sodium chloride. The separated
oily substance was extracted with methylene chloride, washed with
a saturated aqueous solution of sodium chloride, and dried over
anhydrous sodium sulfate. The dried solution was concentrated
under reduced pressure until the amount of methylene chloride
decreased to one half. Isopropyl alcohol was added, and the
mixture was again concentrated under reduced pressure to give a
123~3~L
-28- 7566-720D
colorless solid. The solid was collected by flltration, and
washed with ether to give 2.6 g (yield 51%) of Ampicillin
(5-methyl-2-oxo-1,3-dioxolen-4-yl) methyl ester hydrochloride as
a colorless amorphous solid.
The melting point and spectroscopic data of this product
were identical with those of the product obtained in Example 2 (2).
Example 8
Formulation of an antibacterial agent:-
(1) Ampicillin (5-methyl-2-oxo-1,3-dioxolen-4-yl) methyl
ester hydrochloride 356.7 mg
Lactose 38.3 mg
Magnesium stearate 5.0 mg
400 mgin total
The above ingredients were mixed and encapuslated to
form a capsule.
(2) Ampicillin (5-methyl-2-oxo-1,3-dioxolen-4-yl)methyl ester
hydrochloride 356.7 mg
Lactose 613.3 mg
Hydroxypropyl cellulose 30.0 mg
1,000 mg in total
An ethanol solution of the hydroxypropyl cellulose was
prepared, and added to the Ampicillin ester hydrochloride and
lactose. They were kneaded, extruded through a screen, and dried
to form granules.
(3) Ampicillin (5-methyl-2-oxo-1,3-dioxolen-4-yl) methyl
ester hydrochloride 356.7 mg
~238~
-28a- 7566-720D
Crystalline cellulose 100 mg
Lactose 28.3 mg
Hydroxypropyl cellulose 10 mg
Magnesium stearate 5 mg
500 mg in total
The Ampicillin ester hydrochloride, crystalline
cellulose and lactose were mixed, and an ethanol solution of
hydroxypropyl cellulose was added. They were kneaded and dried.
To the dried mixture was added magnesium stearate. They were
mixed and tabulated to form a tablet.
