JPH034554B2 - - Google Patents

Description

[Detailed description of the invention]

The present invention relates to a novel 7-oxo-1-azabicyclo[3.2.0]hept-2-ene-2-carboxylic acid derivative, more specifically, the following formula: In the formula, Y represents a hydrogen atom or a group -S- _R1 , and _R1 is an unsubstituted or substituted phenyl group,
represents a cyclohexyl group or a 5- or 6-membered aromatic heterocyclic group containing 1 or 2 nitrogen atoms, and R ₂ represents a hydrogen atom or an unsubstituted or substituted benzyl group, and R ₂ is hydrogen This invention relates to a salt of a compound of formula () when an atom is represented, a method for producing the same, and its use as an antibacterial agent. As a 7-oxo-1-azabicyclo[3.2.0]hept-2-ene-2-carboxylic acid derivative, JP-A-64-66697 discloses the following general formula: and the substituents R, R ⁶ at the 2-, 6- and 3-positions,
It gives very broad definitions for R ⁷ and R ⁸ . However, the above-mentioned publication does not disclose the compound of the formula () of the present invention and its analogous compounds at all. The present inventors previously calculated the following formula () We developed a new antibiotic with the structure shown in
This was named antibiotic PS-5 and published [J.
Antibiotics, 31 , 480-482 (1978) and JP-A-Sho.
53-121702 and 1983-30195). The present inventors have conducted repeated research on derivatives that have even better antibacterial activity and/or stability than the above antibiotic PS-5, and have found that the compound represented by the above formula () has extremely excellent antibacterial properties. They discovered this and completed the present invention. In the formula (), the "5- or 6-membered aromatic heterocyclic group containing 1 or 2 nitrogen atoms" includes, for example, 2-pyridyl, 3-pyridyl, 4-pyridyl, 2-pyrimidinyl, 4-pyrimidinyl. , 5-pyrimidinyl, 2-pyrrolyl, 3
-pyrrolyl, 2-imidazolyl, 4-imidazolyl, etc., among which pyridyl and pyrimidinyl groups are preferred. In addition, examples of substituents on the benzene ring in "unsubstituted or substituted phenyl group" and "unsubstituted or substituted benzyl group" include halogen atom, lower alkyl group, lower alkoxy group, lower haloalkyl group, nitro group, lower alkylsulfonyl group, etc. The benzene ring is preferably yo-substituted with such a group. Examples of such "unsubstituted or substituted phenyl groups" include phenyl, p
-nitrophenyl, p-chlorophenyl, p-methoxyphenyl, and the like. On the other hand, specific examples of such "unsubstituted or substituted benzyl group" include benzyl, p-nitrobenzyl, o
-Nitrobenzyl, p-chlorobenzyl, o-chlorobenzyl, p-fluorobenzyl, p-bromobenzyl, p-methoxybenzyl, p-methylsulfonylbenzyl, p-trifluoromethylbenzyl, etc., among which p- The -nitrobenzyl group is advantageous because it can be easily removed by hydrogenolysis. In this specification, the term "lower" means that the group or compound to which this term is attached has 6 or less carbon atoms, preferably 3 or less carbon atoms. Among the compounds represented by the above formula (), a preferred group of compounds is the compound of the formula () where Y represents a group -S-R ₁ and this R ₁ represents the above-mentioned aromatic heterocyclic group. be. Furthermore, in the above formula (), it is more advantageous for R ₂ to be a hydrogen atom than an unsubstituted or substituted benzyl group in terms of antibacterial activity. The compound of formula () when R ₂ is a hydrogen atom can also exist in the form of a salt, examples of such salts include alkali metal salts such as sodium salts, potassium salts, lithium salts; Alkaline earth metal salts such as calcium salts and magnesium salts; other metal salts such as aluminum salts; ammonium salts; monoethylamine, dimethylamine, trimethylamine, monoethanolamine,
Salts with primary, secondary or tertiary amines such as diethanolamine; salts with other organic bases such as benzathine salts, procaine salts, etc. are included, among which pharmaceutically acceptable salts are preferred;
Among these, alkali metal salts such as sodium salts and potassium salts are preferred. Representative examples of the compound of formula () or a salt thereof provided by the present invention are as follows. 3-Cyclohexylthio-6-ethyl-7-oxo-1-azabicyclo[3.2.0]hepta-2-
En-2-carboxylic acid-p-nitrobenzyl ester, 3-cyclohexylthio-6-ethyl-7-oxo-1-azabicyclo[3.2.0]hepta-2-
En-2-carboxylic acid sodium salt, 3-phenylthio-6-ethyl-7-oxo-
1-azabicyclo[3.2.0]hept-2-ene-
2-Carboxylic acid benzyl ester, 3-phenylthio-6-ethyl-7-oxo-
1-azabicyclo[3.2.0]hept-2-ene-
2-carboxylic acid sodium salt, 3-p-nitrophenylthio-6-ethyl-7
-Oxo-1-azabicyclo[3.2.0]hepta-
2-ene-2-carboxylic acid-p-nitrobenzyl ester 3-(4-pyridylthio)-6-ethyl-7-oxo-1-azabicyclo[3.2.0]hepta-2-
En-2-carboxylic acid-p-nitrobenzyl ester, 3-(4-pyridylthio)-6-ethyl-7-oxo-1-azabicyclo[3.2.0]hepta-2-
En-2-carboxylic acid, 3-(2-pyridylthio)-6-ethyl-7-oxo-1-azabicyclo[3.2.0]hepta-2-
En-2-carboxylic acid-benzyl ester, 3-(2-pyridylthio)-6-ethyl-7-oxo-1-azabicyclo[3.2.0]hepta-2-
En-2-carboxylic acid, 3-(2-pyrimidinylthio)-6-ethyl-7
-Oxo-1-azabicyclo[3.2.0]hepta-
2-ene-2-carboxylic acid-p-nitrobenzyl ester, 3-(2-pyrimidinylthio)-6-ethyl-7
-Oxo-1-azabicyclo[3.2.0]hepta-
2-ene-2-carboxylic acid 3-(2-pyrrolylthio)-6-ethyl-7-oxo-1-azabicyclo[3.2.0]hepta-2-
En-2-carboxylic acid benzyl ester, 3-(2-pyrrolylthio)-6-ethyl-7-oxo-1-azabicyclo[3.2.0]hepta-2-
En-2-carboxylic acid, 3-(3-pyridylthio)-6-ethyl-7-oxo-1-azabicyclo[3.2.0]hepta-2-
En-2-carboxylic acid-p-nitrobenzyl ester 3-(3-pyridylthio)-6-ethyl-7-oxo-1-azabicyclo[3.2.0]hepta-2-
En-2-carboxylic acid, 3-(4-pyrimidinylthio)-6-ethyl-7
-Oxo-1-azabicyclo[3.2.0]hepta-
2-ene-2-carboxylic acid-p-nitrobenzyl ester, 3-(4-pyrimidinylthio)-6-ethyl-7
-Oxo-1-azabicyclo[3.2.0]hepta-
2-ene-2-carboxylic acid, 3-(2-imidazolylthio)-6-ethyl-7
-Oxo-1-azabicyclo[3.2.0]hepta-
2-ene-2-carboxylic acid-p-nitrobenzyl ester, 3-(2-imidazolylthio)-6-ethyl-7
-Oxo-1-azabicyclo[3.2.0]hepta-
2-ene-2-carboxylic acid, etc. According to the present invention, Y is a hydrogen atom and R ₂
A compound of the above formula () when is a hydrogen atom,
In other words, the following formula (-c) The compound or its salt represented by the following formula (-
a) In the formula, R ₃ represents a hydrogen atom or an ester residue that can be easily separated by catalytic reduction, or a salt of the compound of formula (-a) when R ₃ is a hydrogen atom is catalytically reduced. It can be manufactured by Catalytic reduction of the compound of formula (-a) or a salt thereof is usually carried out in a suitable solvent inert to the reaction in the presence of a near-neutral buffer, for example in an ether such as dioxane or tetrahydrofuran. This can be carried out by treating the compound of the above formula (-a) or a salt thereof with hydrogen in the presence of a hydrogenation catalyst. Examples of hydrogenation catalysts that can be used include palladium oxide, Raney nickel, etc.
Among them, palladium oxide does not cause a side reaction that saturates the double bond between the 2- and 3-positions,
It is particularly suitable as a catalyst for selectively removing the 3-position side chain. The hydrogen pressure in the above catalytic reduction is not critical, but is generally about 1.0 to about 7.0 Kg/cm ² , preferably about 2.0
It is advantageous to use hydrogen pressures in the range from about 4.0 kg/cm ² to about 4.0 kg/cm 2 and reaction temperatures usually from about 5 to about 50 kg/cm 2 .
°C, preferably within the range of about 20 to about 30 °C. In this way, a compound of formula (-c) or a salt thereof can be obtained in good yield from a compound of formula (-a) or a salt thereof. Isolation of formula (-c) or a salt thereof from the reaction mixture can be carried out by a method known per se. For example, after removing insoluble matter such as a catalyst from the reaction mixture, the liquid can be isolated using QAE Cephadex, QAE cellulose, etc. adsorbed onto a quaternized aminoethyl ion exchange support and eluted with a gradient sodium chloride solution containing a neutral phosphate buffer;
After adding sodium chloride to the solution, it is adsorbed onto a porous styrene-divinylbenzene polymer carrier such as Diamond Ion HP-20AG or Amberlite XAD-2, and after washing with water, it is eluted using the concentration gradient method of water-3% acetone water. A pure compound of formula (-c) or a salt thereof can be obtained by taking a fraction containing the target compound and freeze-drying it. The compound of formula (-a) or its salt used as a starting material in the above method is as described above,
It is a compound known from J. Antibiotic, 31 , 480-482 (1979), JP-A-53-121702 and JP-A-54-30195, etc. Further, as the ester residue R ₃ that can be easily separated by catalytic reduction in formula (-a), for example, substituted or unsubstituted benzyl groups such as benzyl, p-nitrobenzyl, o-nitrobenzyl, etc. are advantageously mentioned. These groups can be obtained by converting the compound of formula (-a) or a salt thereof into a compound of formula R ₃ 'X by esterifying the compound of formula (-a) in a conventional manner.
[wherein _,
- ester-forming functional derivatives such as acid halides and mixed acid anhydrides of the compound of a) are converted into R ₃ 'OH [however,
It can be introduced into the compound of formula (-a) by reacting with an alcohol in which R ₃ ' has the above meaning. The compound of the formula (-c) obtained as described above may be esterified by a conventional method, if necessary, to obtain the compound of the formula (-c), for example, in the same manner as above.
or a salt thereof with an unsubstituted or substituted benzyl halide, or a compound of formula (-c)
By reacting the ester-forming functional derivative of with unsubstituted or substituted benzyl alcohol,
The unsubstituted or substituted benzyl ester can be converted into a compound in which Y represents a hydrogen atom and R ₂ represents an unsubstituted or substituted benzyl ester group in the above formula (). On the other hand, according to the present invention, the compound of the above formula () when Y represents a group -S-R ₁ has the following formula (A): In the formula, R′ ₂ represents an unsubstituted or substituted benzyl group, (i) A compound represented by the following formula () R ₁ −SH () In the formula, R ₁ has the above meaning, and is represented by (B) Then, the following formula (-d) is obtained: In the formula, R ₁ and R ₂ ' have the above-mentioned meanings. The compound represented by the formula can be easily produced by subjecting it to hydrogenolysis, if necessary. The reaction between a compound of formula () and a thiol compound of formula () or a reactive derivative thereof is usually N,N-
In solvents such as dimethylformamide (DMF), tetrahydrofuran (THF), dioxane, hexamethylphosphoramide (HMPA), glyme, etc., especially
In DMF, about -20°C or less, preferably about -30°~
Advantageously, it is carried out under low temperature conditions in the range of about -50°C. The reaction can be completed under such conditions, usually within 15 minutes to 60 minutes. When the thiol compound of formula () is used as it is, it is preferable to carry out the reaction in the presence of a base, and usable bases include, for example, sodium hydride, potassium hydride, sodium amide, potassium amide, hydroxide. Sodium, potassium hydroxide, sodium ethoxide, sodium methoxide, potassium butoxide, triethylamine, tripropylamine, etc. are included, and these bases are used in an amount of at least 0.9 equivalents, preferably 1.0 to 1.2 equivalents, per mole of the thiol compound. It is advantageous to use it in proportions. Furthermore, in the above reaction, instead of the presence of a base, a reactive derivative of the thiol compound of formula () may be used, and examples of such reactive derivatives include sodium ethanethiolate, sodium butanethiolate, potassium ethanethiolate,
Potassium butane thiolate and the like can be mentioned. Although the amount of the thiol compound of formula () or its reactive derivative used is not critical, it is generally used in a ratio of at least 1.0 mol, preferably 1.1 to 1.5 mol, per 1 mol of the compound of formula (). It's advantageous. In this way, a compound of the above formula (-d) is obtained, which can be isolated and purified by the following method if necessary, and then subjected to hydrogenolysis to remove the group R ₂ '. This allows the following formula (-e) In the formula, R ₁ has the above-mentioned meaning and can be changed to a compound represented by the following or a salt thereof. Isolation and purification of the compound of the above formula (-d) can be carried out, for example, by diluting the reaction mixture with an inert solvent that is immiscible with water, such as benzene, toluene, methylene chloride, ethyl acetate, etc., and diluting the reaction mixture with a PH8.7 phosphate buffer. liquid or sodium bicarbonate solution and then PH6.8
After washing with phosphate buffer and drying this solvent layer over anhydrous sodium sulfate, it was evaporated under reduced pressure, and the residue was dissolved in a small amount of the above inert solvent. This can be carried out by passing the reaction mixture through a column (manufactured by BioRad) and developing with the same solvent, or, if necessary, by a combination of methods such as passing through a silica gel column. compounds can be isolated. Hydrogenolysis of the compound of formula (d) can be carried out according to a method known per se, for example, in a solvent such as water:ethers such as dioxane and tetrahydrofuran, using a near-neutral buffer. This can be carried out by treatment with hydrogen in the presence of a hydrogenation catalyst such as platinum oxide, palladium on carbon, or palladium black. The compound of formula (-e) thus obtained can be isolated from the reaction mixture, usually in the form of a salt, by the method described above. The compound of formula () used as a starting material in the above method is a new compound, for example the antibiotic PS-
5, by an esterification method similar to that used for conventionally known carboxyl group-containing antibiotics (e.g. penicillin, cephalosporin, etc.).
For example, by the esterification method described in lines 36-48 of JP-A-52-149374, the following formula () In the formula, R ₂ ' has the above-mentioned meaning, and can be produced by forming an unsubstituted or substituted benzyl ester of antibiotic PS-5 represented by and then S-oxidizing the compound of formula (). can. The S-oxidation of the compound of formula () can be carried out by methods known per se, for example, the S-oxidation of sulfur-containing β-lactam antibiotics such as penicillins and cephalosporins.
The oxidation can be carried out according to methods often used for oxidation. For example, the compound of the formula () is a mild oxidizing agent that does not substantially act on the 7-oxo-1-azabicyclo[3.2.0]hept-2-ene-2-carboxylic acid skeleton, that is, perbenzoic acid, It can be reacted with ozone, phenidichloroiodide, hydrogen peroxide, selenium dioxide, or sodium metaperiodate. Suitable such mild oxidizing agents are organic peracids, in particular perbenzoic acid and m-chloroperbenzoic acid, most preferably substituted perbenzoic acids such as m-chloroperbenzoic acid. . The reaction between the compound of formula () and the above oxidizing agent can be carried out using an inert solvent such as methylene chloride, chloroform,
in a solvent such as carbon tetrachloride at room temperature or lower;
It is advantageous to conduct the reaction under mild temperature conditions, preferably about -30° to about 20°C. The reaction can be completed under such conditions, usually within 3 minutes to 3 hours. The amount of the oxidizing agent used to oxidize the compound of formula () can vary widely depending on the type of oxidizing agent, reaction conditions, etc.;
A range of 0.3 to 1.8 molar equivalents, preferably 0.7 to 1.3 molar equivalents per mole of the compound is useful. After completion of the reaction, the compound of formula (), which is the S-oxidation product, can be isolated and purified by various methods known per se. It can be isolated from the reaction solution depending on the method utilized. The compound of the formula () obtained by the method described above, in which R ₂ is a hydrogen atom, can be prepared by methods known per se, such as sodium hydrogen carbonate, disodium phosphate, dipotassium phosphate, 2-ethyl It can be converted into a salt by treatment with an inorganic base such as potassium hexanoate or ammonia; or an organic base such as monoethylamine, dimethylamine, triethylamine, monoethanolamine, diethanolamine, benzathine, or procaine. The compound of formula () used as starting material in the above method, e.g. antibiotic PS-5, is
J. Antibiotics, 31 , 480-482 (1978) and JP-A-58-121702 and JP-A-54-30195, antibiotics obtained by fermentation are produced by fermentation. PS-5 is the following formula (-b) It has been confirmed that the compound of the above formula () of the present invention derived from this antibiotic PS-5 also has the following formula (-a). In the formula, Y and R ₂ have the meanings given above, and have a 5,6-trans configuration. Compounds of the above formula () provided by the present invention, especially the following formula (-b) In the formula, Y has the above-mentioned meaning. The compound or a salt thereof has very excellent antibacterial activity. For example, the following formula The antibacterial activity of the compound represented by or its sodium salt is as shown in Table 1 below.

【table】

[Table] The original names of the test bacteria used in Table 1 above are as follows. Bacillus subtilis ATCC6633
ATCC6633) Sarcina Lutea
lutea) Staphylococcus aureus FDA209P (Staphylococcus
aureus FAD209P) Staphylococcus aureus Smith strain (Staphylococcus aureus FAD209P)
Staphylococcus aureus Smith)
aureus Russell) Staphylococcus epidermidis
epidermidis) Alcaligenes fuecalis A1 (Alcaligenes epidermidis)
faecalis A1) Citrobacter freundei GN346
(Citrobacter freundii GN346) Comamonas terrigena B-996 (Comamonas
terrigena B-996) Enterobacter aerogenes E19
(Enterobacter aerogenes E19) Enterobacter cloacae 45
(Enterobacter cloacae 45) Enterobacter E8 (Enterobacter sp E8) Escherichia coli K-12 RGN823 (Escherichia coli RGN823) Klebsiella pneumoniae K13
pneumoniae K-13) Proteus mirabilis P6 (Proteus mirabilis P6)
mirabillis P6) 〃 ・Proteus rettgeri P7 (Proteus rettgeri
P7) 〃 ・Proteus vulgaris GN (Proteus vulgaris)
GN76) 〃 P22 (Proteus sp. P22) Providencia P8 (Providencia sp.P8) Pseudomonas aeruginosa IFO3445 (Pseudomonas aeruginosa
IFO3445) Pseudomonas aeruginosa NCTC10490 (Pseudomonas aeruginosa)
NCTC10490) Serratia marcescens S18 (Serratia
marcescens S18) T55 (Serratia marcescens T55) The above antibacterial activity was measured as follows. Namely, heart infusion agar medium “Difco” [manufactured by Difco Laboratories]
The minimum inhibitory concentration of the compound of the formula (-b) against the various test bacteria in Table 1 above was determined by the agar dilution assay using the above. The specimen was dissolved in sterile distilled water, and multiple dilutions were prepared. After dissolving and sterilizing 1 ml of this antibiotic solution, approx.
The mixture was mixed with 9 ml of Heart Infusion Agar medium (Difco) heated to 60°C in a 9 cm diameter shear dish and made into a flat plate. Trypto soy broth medium [manufactured by Eiken Chemical Co., Ltd.]
The inoculum of the test bacteria shown in the table above obtained by static culture at 35°C for 18 to 20 hours was diluted with the same trypto soy broth medium to adjust the number of bacteria to approximately 10 ⁸ cells/ml. Agar plates containing the antibiotic were inoculated using a multi-inoculator. After culturing this plate at 35°C for 20 hours, the presence or absence of growth is observed, and the lowest concentration at which no growth is observed is defined as the minimum inhibitory concentration of the compound against the bacteria. The characteristics of the antibacterial spectrum of the compound of formula (-b) thus measured are as follows. The PS-5 sodium salt, which is the starting material of the derived compound of formula (-b), exhibits superior antibacterial activity compared to cefazolin (CEZ), which is commonly used among known β-lactam drugs, and has been shown to be useful. It is known to be effective against Pseudomonas aeruginosa, Serratia sp., Proteus sp., Klebsiella sp. and Enterobacter sp.
However, the compound of formula (-b) has more general antibacterial activity than PS-5 sodium salt, which has excellent antibacterial activity, and especially 3-heterocyclylthio derivatives having an aromatic heterocyclic group. has about 10 to 20 times more antibacterial activity against Gram-positive bacteria, and has the same or higher activity against Gram-negative bacteria, for example, about 2 to 5 times more activity. In particular, the 3-(4-pyridylthio) derivative (compound 1) can be cited as the most antibacterial compound. In addition, 2 mg of the compound of formula (-b) was administered subcutaneously to each of 5 mice, and 5 minutes after administration, a small amount of blood was collected from the eye into a heparinized capillary tube and centrifuged. The concentration of the compound in the supernatant (plasma, plasma) was measured by a bioassay method using Comamonasterigena B-996 as a test bacterium. Table 2 below shows the average values of 5 mice.

Table: All compounds of formula (-b) tested were well absorbed and gave fairly high maximum concentrations. Among them, 3-
Heterocyclylthio-derivatives gave excellent highest concentrations. In addition, a compound having an aromatic group in the S side chain at the 3-position in formula (-b) had a long half-life and good persistence of blood concentration. Next, mice were intraperitoneally infected with Staphylocotcus aureus Smith strain at 5 × 10 ⁵ cells per mouse (5 mice per group, male ddy mice, Shizuoka).
The in vivo activity of the compound of formula (-b) was measured using 2 hours after infection, expression (-b)
An injection containing the sodium salt of the compound was administered subcutaneously. The results ( _CD50 ) of the infection treatment test are shown in Table 3 below. PS-5 sodium salt of raw material

[Table] As is clear from the results of this Table-3, 4-
The pyridylthio compound (compound 1), 2-pyridylthio compound (compound 2) and 2-pyrimidinylthio compound (compound 3) are
5. Shows a clearly superior therapeutic effect than the sodium salt. Furthermore, among the compounds of formula (-b), 4-pyridylthio compound (compound 1), 2-pyridylthio compound (compound 2), 2-pyrimidinylthio compound (compound 3), and 3-phenylthio compound (compound 5), Although up to 20 mg (1 g/Kg) per mouse was administered intraperitoneally, no compound was observed that caused mouse death. From the above in vitro and in vivo activity data, the following formula (-b) In the compound or its salt represented by, Y=-S
- Compounds where R ₁ is a nitrogen-containing heterocyclic group, that is, compounds where _{R 1 is} 4-pyridyl, 2-bilidyl, 2-pyrimidyl, etc. (the above compounds 1, 2 and 3 etc.) and their salts are particularly suitable as antibacterial agents. As mentioned above, the compound of formula (-b) or a salt thereof exhibits antibacterial activity and can be used as an active ingredient of an antibacterial agent for the prevention, treatment and/or treatment of infectious diseases caused by Gram-positive and Gram-negative bacteria. It can be effectively used for the prevention, treatment, treatment, etc. of bacterial infections not only for humans but also for non-human animals such as mammals, poultry, and fish. The compound of the formula (-b) or a salt thereof can be administered orally, topically, or parenterally (intravenously, intramuscularly, intraperitoneally, etc.), and depending on the method of administration, the usual procedure is as follows. It can be formulated and used in various drug forms as described above. For example, the compound of formula (-b) or a salt thereof may be present in a solid form (e.g., tablet, capsule, powder, granule, dragee, troche, powder, spray) together with a pharmaceutically acceptable carrier material, diluent, etc. , suppositories, etc.), semisolid forms (e.g., ointments, creams, semisolid capsules, etc.), or liquid forms (e.g., solutions, emulsions, suspensions, lotions, syrups, injections, liquid sprays, etc.). can do. The unit dosage form containing the compound of formula (-b) or a salt thereof, whether liquid, semi-solid or solid, generally contains 0.1 to 99% by weight, preferably 10% by weight.
May contain ~60% by weight of active ingredient.
The content of the active ingredient in the drug varies depending on the dosage form and total amount of the drug, but is usually about 10 mg to about 1000 mg.
mg, preferably in the range of about 100 mg to about 1000 mg. The unit dosage form for parenteral administration is usually a compound of formula (-b) of the present invention or a salt thereof dissolved in sterile water with a purity close to 100%, or a compound with a solubility that can be easily made into a solution. It can be made into powder. Next, the present invention will be further explained by examples. Example 1 3-(2-acetamidoethylthio)-6-ethyl-7-oxo-1-azabicyclo[3.2.0]hept-2-ene-2-carboxylic acid benzyl ester (PS-5.benzyl ester 3-(2-acetamidoethylthio) produced by the method described in J. Antibiotics, 31 , 480-482 (1978)
Sodium -6-ethyl-7-oxo-1-azabicyclo[3.2.0]hept-2-ene-2-carboxylate (hereinafter abbreviated as PS-5 sodium salt)
4.75 ml of triethylamine was added to a 200 ml solution of 1204 mg (purity 82%) of dry dimethylformamide (hereinafter abbreviated as DMF), and 3.68 ml of benzyl bromide was added while stirring at -70°C in a nitrogen stream.
After returning to room temperature and stirring for 3 hours, the reaction mixture was
After pouring into 1000 ml of benzene and washing three times with 300 ml of 0.1M, PH6.8 phosphate buffer, the solvent layer was dried over anhydrous sodium sulfate. After distilling off the solvent under reduced pressure at a concentration of 30% or less, it was dissolved in a small amount of benzene and subjected to column chromatography (120 g, 3.2 x 75.0 cm) using Bio-Bee S-X3 with benzene as a developing agent. Next, each eluted fraction was confirmed by thin layer chromatography (hereinafter abbreviated as TLC), and fractions containing the target compound were collected.The solvent was distilled off under reduced pressure to obtain 1081 mg of a colorless oil. [α] ²¹ _D +19.97° (c0.33, CHCl ₃ ). UVλ ^MeOH _nax nm (ε): 318 (11000). IRν ^CHCl3 _nax cm ^-1 : 3430 (CO NH ), 1770 (β-lactam
CO), 1690 (ester CO), 1665 ( CO NH). NMR (CDCl ₃ ) δ: 1.04 (3H, t, J = 8Hz,
CH ₂ CH ₃ ), 1.70-2.00 (2H, m, CH ₂ CH ₃ ), 1.96
(3H, s, COCH ₃ ), 2.80~3.60 (7H, m,
3XCH ₂ , CH), 3.94 (1H, dt, J=8Hz, J=3
Hz, C-5H), 5.21 (1H, d, J=12Hz, ArC
H・H), 5.38 (1H, d, J=12Hz, ArCH・H ),
5.90~6.10 (1H, brs, NH ), 7.34 (5H,
distorted, s, Ar H ). Mass (m/e): 388 (M ⁺ ). Example 2 3-(2-acetamidoethylthio)-6-ethyl-7-oxo-1-azabicyclo[3.2.0]hept-2-ene-2-carboxylic acid-(p-nitrobenzyl) ester ( Preparation of PS-5 p-nitrobenzyl ester) 275 mg of PS-5 sodium salt was dissolved in 20 ml of dry DMF, and triethylamine was added while stirring at room temperature.
Added 197μ. After sufficient time, a solution of 179 mg of p-nitrobenzyl bromide dissolved in 5 ml of dry DMF was added at 5° C. with stirring, and the mixture was stirred at the same temperature for 3 hours. After the reaction is complete, the reaction mixture is diluted with benzyl 150
ml and washed three times with 100 ml of 0.1M, PH6.8 phosphate buffer. After drying over anhydrous sodium sulfate, the solution was distilled off under reduced pressure, and the residue was poured into 30 g of silica gel.
Column chromatography using (1.8 x 30 cm)
Yellow crystals were obtained as a solvent distillation residue from the benzene-acetone (3:1) elution section. This was recrystallized from benzene to obtain 240 mg of yellow needle crystals with a mp of 163-165°C. [α] ²¹ _D +70.7° (c1.00, CHCl ₃ ). UVλ ^CHCl3 _nax nm (ε): 322 (12800), 269 (12000). IRν ^CHCl3 _nax cm ^-1 : 3460 ( NH CO), 1780 (β-lactam
CO), 1710 (ester CO), 1675 ( NHCO ), 1560
( _NO2 ), 1350 ( _NO2 ). NMR (CDCl ₃ ) δ: 1.04 (3H, t, J = 8Hz,
CH ₂ CH ₃ ), 1.90 (2H, dq, J=8Hz, J=2Hz,
CH ₂ CH ₃ ), 1.96 (3H, s, NHCOC H ₃ ), 2.80~
3.60 (7H, m, 3XCH ₂ , CH), 3.98 (1H, dt, J
=8Hz, J=3Hz, C- 5H ), 5.19(1H, d,
J=14Hz, ArC H・H), 5.49 (1H, d, J=14
Hz, ArCH・H ), 5.95 (1H, brs, NH ), 7.61
(2H, d, J = 9Hz, Ar H ), 8.18 (2H, d, J
=9Hz, ArH ). Mass (m/e): 433 (M ⁺ ), 363 (M ⁺ −EtCH=C
=O). A na l.Calcd.for C ₂₀ H ₂₃ O ₆ N ₃ S:C, 55.43:H,
5.35:N, 9.70. Found:C, 55.29:H, 5.24:
N, 9.40. Example 3 Process for producing sulfoxide of PS-5. benzyl ester 50 mg of PS-5. benzyl ester obtained in Example 1
Dissolve in 5 ml of dry methylene chloride, cool with ice water,
While stirring, add 28.74 mg of m-chloroperbenzoic acid.
(1.1 times the mole relative to the raw material) in 1 ml of dry methylene chloride
A solution dissolved in was added dropwise to the mixture, and the mixture was vigorously stirred at the same temperature for 15 minutes. After the reaction is completed, dilute with 50 ml of methylene chloride and wash twice with 25 ml each of 0.1M, PH8.7 and 6.8 phosphate buffers. The solvent layer was dried over anhydrous sodium sulfate, evaporated under reduced pressure, and immediately dissolved in 1 ml of benzene. Bio-Beas S-X3 column (1.2 x 75.0cm) pre-swollen with benzene
The concentrated solution was added to the solution, and after developing with benzene, fractions containing the target substance were collected, and the solvent was distilled off under reduced pressure to obtain 53.5 mg of a colorless dry solid. [α] ²¹ _D −43.96 (c1.00, MeOH). UVλ ^MeOH _nax nm (ε): 307 (4440). IRv ^CHCl3 _nax cm ^-1 : 3400 ( NHCO ), 1790 (β-lactam CO), 1710 (ester CO), 1670 ( NHCO ). NMR (CDCl ₃ ) δ: 1.04 (8H, t, J = 8Hz, J
= _3H2CH2CH3 ), 1.64~ _2.02 (2H _, m, _CH2
CH3 ), 1.94 (3H, s, _COCH3 ), 2.88-3.80 (7H,
m, 3XCH ₂ , C H ), 3.88~4.20 (1H, m, C
−5 H ), 5.26 (2H, s, ArC H ₂ ), 7.36 (5H,
s, ArH ). Mass (m/e): 404 (M ⁺ ). Example 4 Preparation of sulfoxide of PS-5.p-nitrobenzyl ester 240 mg of PS-5.p-nitrobenzyl ester was dissolved in 25 ml of dry methylene chloride and cooled to -30°C. 104.9 mg of m-chloroperbenzoic acid was dissolved in 12 ml of dry methylene chloride, added to the above solution with stirring, and reacted at the same temperature for 40 minutes. After the reaction is complete, add triethylamine to 100ml of ethyl acetate in advance.
An ice-cooled solution was prepared by adding 0.09 ml, and the reaction solution was poured into this solution. After cooling on ice and stirring for 5 minutes, add 100 ml of ethyl acetate and transfer to a separatory funnel.
Washed four times with ice-cold 0.1 MPH 6.8 phosphate buffer.
After drying the solvent layer over anhydrous sodium sulfate, benzene was added and the mixture was concentrated under reduced pressure to 1 ml. The concentrated solution was passed through a 18 g silica gel column (42 ml, 2.0 x 13.4 cm, same as the above silica gel) pre-wetted with benzene-acetone (1:1), and developed with about 50 ml of the same solvent. 14.1mg of raw material was eluted.
Further, the title target compound was eluted (yield
211mg, 89%). The Rf value of this material on a silica gel TLC plate (same as the above TLC plate) was 0.36 in benzene-acetone (1:2). [α] ²¹ _D +18.0° (c1.00, CHCl ₃ ). UVλ ^CHCl3 _nax nm (ε): 267 (13079), 310 (7775). IRν ^CHCl3 _nax cm ^-1 : 3460 ( NH CO), 1785 (β-lactam
CO), 1720 (ester CO), 1680 ( NHCO ). NMR ( _CDCl3 ) δ: 1.00 (3H, t, J= 7.0Hz, _{CH2CH3 )} , 1.70~
2.00 (2H, m, CH ₂ CH ₃ ), 2.00 (3H, s,
NHCO CH ₃ ), 3.00~3.88 (7H, m, 3XCH ₂ ,
CH), 3.92~4.28 (1H, m, CH), 5.12~5.60 (2H, distorted d, ArC H・H,
ArCH.H ), 6.25-6.55 (1H, brs, NH ), 7.52-7.72 (2H, ArH ), 8.12-8.28 (2H, ArH ). Mass (m/e): 449 (M ⁺ )FD Mass). Example 5 3-cyclohexylthio-6-ethyl-7-oxo-1-azabicyclo[3.2.0]hepta-2
-Preparation of -ene-2-carboxylic acid-p-nitrobenzyl ester PS-5.To a solution of 300 mg of sulfoxide of p-nitrobenzyl ester in 40 ml of anhydrous DMF was cooled to -45°C, and while stirring, cyclohexanethiol was added.
Add 106μ and triethylamine 111μ,
After continuing to stir at the same temperature for 40 minutes, the reaction mixture was poured into 200 ml of benzene, washed with 4 x 150 ml of 0.1M PH6.8 phosphate buffer, and the combined aqueous layer was poured with 70 ml of benzene 3 x. The extracted and combined organic layers were further washed with 70 ml of the same buffer twice, dried over anhydrous sodium sulfate, and concentrated under reduced pressure. The obtained yellow oil was subjected to silica gel column chromatography (30
g, 2.6 x 12 cm, solvent system benzene-acetone =
40:1) to obtain 175.2 mg of the target yellow powder.
was obtained (yield 61%). Rf: 0.43 (benzene-acetone = 20:1). [α] ²² _D : +42.6 (c1.00, THF). UVλ ^THF _nax nm (ε): 325 (15600), 270 (12800). IRν ^CHCl3 _nax cm ^-1 : 1770, (β-lactam CO), 1695
(Ester CO)). NMR ( _CDCl3 ) δ: 1.08 (3H, t, J=7.5Hz _{, CH2CH3} ), 1.2-2.1 (12H, m _{, CH2CH3} ), Cyclohexyl
CH ₂ ), 2.8-3.2 (4H, m, C-4H, C-6H,
SCH), 3.94 (1H, dt, J=9.0Hz, J=3.0 PH, C-
5H), 5.18 (1H, d, J=14Hz, ArC H・H), 5.50 (1H, d, J=14Hz, ArCH・H ), 7.62 (2H, d, J=9.0Hz, ArH), 8.18 ( 2H, d, J = 9.0Hz, ArH). Mass (m/e): 430 (M ⁺ ), 360 (M ⁺ −Et−CH=
C=O) Example 6 3-cyclohexylthio-6-ethyl-7-oxo-1-azabicyclo[3.2.0]hepta-2
-Manufacture of ene-2-carboxylic acid/sodium salt 3-cyclohexylthio-6-ethyl-7-oxo-1-azabicyclo[3.2.0]hepta-2-
37 mg of ene-2-carboxylic acid p-nitrobenzyl ester was dissolved in a mixed solvent of 2 ml of dioxane and 1.5 ml of 0.1 M, PH8.4 phosphate buffer, 40 mg of platinum oxide was added, and a Parr reduction apparatus (hydrogen Pressure 4Kg
The reaction was carried out for 4 hours at room temperature (r/cm ² ). Afterwards, add a supernatant to remove the catalyst, and pre-prepare the filtrate.
QAE-Sephadex A-25 column (1.1 x 20 cm: quaternized aminoethyl dextran gel, manufactured by Pharmacia) treated with 0.01M phosphate buffer
0 to 0.4 M sodium chloride concentration gradient method (100 ml of 0.01 M phosphate buffer,
0.01M phosphate buffer with 0.4M sodium chloride concentration
(100 ml was used). In this eluate, λ _nax
301 _on , collect the ultraviolet absorbing categories, add sodium chloride to make the salt concentration 3%, and add diamond ion.
Adsorb onto HP-20AG column (1.1 x 20 cm, porous divinylbenzene-styrene polymer: manufactured by Mitsubishi Kasei Corporation), and use 0-30% acetone gradient gradient method (using 100 ml of ion-exchanged water and 100 ml of 30% acetone). The fraction having ultraviolet absorption at λ _nax 301 _on was collected in a high-performance liquid chromatogram and lyophilized to obtain 90 mg of the target product (yield 33%). UVλ ^H2O* _nax nm (ε): 305 (6600). *0.1M phosphate buffer (PH
_7.0 ) NMR (D2O) δ: 1.04 (3H, t, J=7.5Hz, _{CH2CH3 ) ,} 1.2-2.2 (12H, m, _CH2CH3 , cyclohexyl
_CH2 ) 3.0~3.4 (4H, m, C-4H, C-6H,
S- CH ), 4.02 (1H, dt, J=9Hz, J=3Hz C-5
H). Example 7 Preparation of 3-(phenylthio)-6-ethyl-7-oxo-1-azabicyclo[3.2.0]hept-2-ene-2-carboxylic acid-p-nitrobenzyl ester 100 mg (0.223 mmol) of PS -5.p-Nitrobenzyl ester sulfoxide in 80 ml of anhydrous
Dissolved in DMF and cooled to -50°C. 37 μ (0.267 mmol) of triethylamine was added to this solution while stirring. Furthermore, 27.4 μ (0.267 mmol) of thiophenol was added and the mixture was allowed to react for 10 minutes. The reaction solution was poured into 300ml of benzene, washed with 0.1M phosphate buffer (Hz6.8) (150ml x 3), and the organic layer was dried over anhydrous sodium sulfate.The benzene was distilled off under reduced pressure and the A yellow oil was obtained. This product was applied to a column (φ1.2×20 cm) filled with 10 g of silica gel and benzene, and developed and eluted with a solvent of benzene-acetone (80:1v/v).
After spotting a part of it on thin layer chromatography and developing it, irradiating it with 360 nm UV light to search for the target object, collecting the sections containing the target object, and distilling off the solvent under reduced pressure, 55.1 mg of the target product was obtained (yield 65.2%). [α] ²² _D +3.0° (c1.00, THF). UVλ ^THF _nax nm (ε): 320 (11000), 271 (9100). IRν ^CHCl3 _nax cm ^-1 : 1780 (β-lactam CO), 1710 (ester CO). NMR ( _CDCl3 ) δ: 1.00 (3H, t, J=7, 5Hz, _CH2CH3 ), 1.6-1.90 (2H, m _{, CH2CH3} ), 2.66 (2H, _d , J=9Hz, C -4 H), 3.02 (1H, dt, J=9Hz, J=4Hz, C-6
H), 3.82 (1H, dt, J=9Hz, J=4Hz, C-5
H), 5.24, 5.54 (2H, each d, J=14Hz, ArC H
H), 7.28-7.60 (5H, m, ArH ), 7.68 (2H, d, J=9Hz, ArH), 8.24 (2H, d, J=9Hz, ArH). Mass (m/e): 424 (M ⁺ ), 354 (M ⁺ −EtCH=C=
O). Example 8 3(phenylthio)-6-ethyl-7-oxo-
1-Azabicyclo[3.2.0]hept-2-ene-2-carboxylic acid sodium salt 3-phenylthio-6-ethyl-7-oxo-
1-azabicyclo[3.2.0]hept-2-ene-
2-Carboxylic acid p-nitrobenzyl ester 120
mg to dioxane 6.5ml−0.1M phosphate buffer (PH
8.5) Dissolved in 6.0 ml of mixed solvent, added 100 mg of platinum oxide, and reacted for 4 hours at room temperature in a Parr reduction apparatus (hydrogen pressure 4 Kg/cm ² ). Separate the catalyst
The solution was treated and purified in the same manner as in Example 6 to obtain 31.8 mg of the desired product (yield 36%). UVλ ^H2O* _nax nm (ε): 303 (10600). *0.1M phosphate buffer NMR ( _D2O ) δ: 0.96 (3H, t, J=8Hz, _CH2CH3 ), 1.60-1.90 (2H _, m _{, CH2CH3} ), 2.40-2.90 ( 2H, m, C-4 H), 3.15 (1H, dt, J=9Hz, J=3Hz, C-6
H) 3.86 (1H, dt, J=9Hz, J=3Hz, C-5
H), 7.35-7.65 (5H, m, ArH ). Example 9 3-(p-nitrophenylthio)-6-ethyl-
Preparation of 7-oxo-1-azabicyclo[3.2.0]hept-2-ene-2-carboxylic acid p-nitrobenzyl ester Add 200 mg (0.445 mmol) of the sulfoxide of PS-5 p-nitrobenzyl ester to 15 ml of anhydrous DMF. After melting and cooling to -45℃ and adding 160μ (1.15mmol) of triethylamine while stirring, add 138mg (0.890mmol) of p-nitrothiophenol to anhydrous solution.
The mixture was dissolved in 0.5 ml of DMF, added, and reacted for 60 minutes.
Thereafter, the product was purified in the same manner as in Example 7 to obtain 51.8 mg of the target product (yield 24.8%). [α] ²² _D +32.2° (c1.00, THF). UVλ ^THF _nax nm (ε): 266 (17800), 310sh (12400). IRν ^CHCl3 _nax cm ^-1 : 1755 (β-lactam CO), 1700 (ester CO). NMR ( _CDCl3 ) δ: 1.02 (3H, t, J = 7.5Hz, _CH2CH3 ), 1.60-2.00 (2H, m _{, CH2CH3} ), 2.78 (2H, _d , J = 9.0Hz, C -4 H), 2.96~3.20 (1H, m, C-6 H), 3.92 (1H, dt, J=9Hz, J=3Hz, C-5
H), 5.24 (1H, d, J=14Hz, ArC H・H), 5.50 (1H, d, J=14Hz, ArCH・H ), 7.64 (4H, d, J=8Hz, ArH), 8.18 (4H , d, J=8Hz, ArH). Mass (m/e): 469 (M ⁺ ), 399 (M ⁺ −Et−CH=C=
O). Example 10 Production of 3-(phenylthio)-6-ethyl-7-oxo-1-azabicyclo[3.2.0]hept-2-ene-2-carboxylic acid benzyl ester In Example 7, PS-5.p - Sulfoxide of nitrobenzyl ester 100 mg (0.223 mmol)
Purification was carried out in the same reaction and treatment method as in Example 7, except that 86 mg (0.220 mmol) of PS-5 benzyl ester sulfoxide was used instead of, to obtain 52.0 mg of the target product (yield 71.9%). . UVλ ^THF _nax nm (ε): 319 (11500). IRν ^CHCl3 _nax cm ^-1 : 1775 (β-lactam CO), 1700 (ester CO). NMR ( _CDCl3 ) δ: 0.96 (3H, t, J=7.5Hz, _CH2CH3 ), 1.60-1.90 (2H, m _{, CH2CH3} ), 2.62 (2H, _d , J=9Hz, C- 4 H ₂ ), 2.98 (1H, dt, J=9Hz, J=4Hz, C-6
H), 3.78 (1H, dt, J=9Hz, J=4Hz, C-5
H), 5.27 (2H, d, J=12Hz, ArCH.H ), 5.42 (2H, d, J=12Hz, ArCH.H), 7.20-7.60 (10H, m, ArH). Mass (m/e): 379 (M ⁺ ), 309 (M ⁺ −EtCH=C=
O). Example 11 3-(4-pyridylthio)-6-ethyl-7-oxo-1-azabicyclo[3.2.0]hepta-2
Preparation of -ene-2-carboxylic acid p-nitrobenzyl ester 150 mg (0.372 mmol) of PS-5 p-nitrobenzyl sulfoxide was dissolved in 40 ml of dry DMF,
Cooled to -50°C. Add to this solution while stirring.
A solution of 64.5 mg (0.488 mmol) of the sodium salt of 4-pyridylmercaptan dissolved in 10 ml of dry DMF was added and reacted for 40 minutes. 150ml of reaction solution
of benzene, and this was washed with 0.1M phosphate buffer (PH8.5) (70ml x 3). The organic layer was dried over anhydrous sodium sulfate, and benzene was distilled off under reduced pressure to obtain a pale yellow oil. This was applied to a column (1.0 x 15 cm) filled with 8 g of silica gel and benzene.
It was developed and eluted with acetone (5:1v/v) solvent.
The target product was searched for from each fraction by thin layer chromatography, the fractions containing the target product were collected, and the solvent was concentrated under reduced pressure to obtain 47mg of the target product.
The desired product was obtained (yield 29.7%). [α] ²² _D : +18.4° (C1.00, THF). UVλ ^THF _nax nm (ε): 322 (7900), 267 (8700). IR ^CHCl3 _nax cm ^-1 : 1785 (β-lactam CO), 1720 (ester CO). NMR ( _CDCl3 ) δ: 1.10 (3H, t, J=7., 5Hz _{, CH2CH3} ), 1.64-2.04 (2H, m _{, CH2CH3} ) 2.60-3.24 (3H, m, C-4 H, C-6
H), 3.92 (1H, dt, J=9Hz, J=3Hz, C-5
H), 5.30 (1H, d, J=14Hz, ArC H・H), 5.58 (1H, d, J=14Hz, ArCH・H ), 7.40 (2H, dd, J=7Hz, J=3Hz), PyH ), 7.68 (2H, d, J=9Hz, ArH), 8.41 (2H, d, J=9Hz, ArH), 8.60 (2H, dd, J=7Hz, J=8Hz, PyH). Mass (m/e): 425 (M ⁺ ), 355 (M ⁺ -EtCH=C=O). Example 12 3-(4-pyridylthio)-6-ethyl-7-oxo-1-azabicyclo[3.2.0]hepta-2
Production of -ene-2-carboxylic acid 3-(4-pyridylthio)-6-ethyl-7-oxo-1-azabicyclo[3.2.0]hepta-2-
p-nitrobenzyl ene-2-carboxylate 54 mg
was dissolved in a mixed solvent of 2 ml of dioxane and 2 ml of 0.1 M phosphate buffer (PH 7.0), 54 mg of platinum oxide was added, and the mixture was heated in a Parr reduction apparatus (hydrogen pressure 4 Kg/cm ² ).
The reaction was carried out at room temperature for 4 hours. Separate the catalyst
The liquid and washing liquid were combined and concentrated under reduced pressure to about two-thirds of the volume. 2.5 g of sodium chloride was added to this liquid, and water was further added to make 50 ml. This is a diamond ion
It was applied to a HP20AG column (φ1.1 x 20 cm, same as the above resin) and eluted by 0-30% acetone water gradient gradient method with a total volume of 300 ml. The fractions were searched by high performance liquid chromatography, and fractions having ultraviolet absorption at λ _nax 303 nm were collected and freeze-dried to obtain 20.7 mg of the target product (yield 56.3%). UVλ ^H2O* _nax nm (ε): 303 (8500). *0.1M phosphate buffer (PH7.0) NMR (D2O) δ: 1.00 (3H, t, J=7Hz, _{CH2CH3 ) , 1.64-2.00} (2H, m, _CH2CH3 ), 2.92 (2H, d, J=9Hz, C-4 H), 3.34 (1H, dt, J=9Hz, J=3Hz, C-6
H), 4.06 (1H, dt, J=9Hz, J=3Hz, C-5
H), 7.50 (2H, dd, J=7Hz, J=8Hz, PyH), 8.30-8.60 (2H, m, PyH). Example 13 3-(4-pyridylthio)-6-ethyl-7-oxo-1-azabicyclo[3.2.0]hepta-2
-Manufacture of -ene-2-carboxylic acid-benzyl ester In Example 11, 150 mg (0.372 mol) of sulfoxide of PS-5 p-nitrobenzyl ester
Example except that 43 mg (0.124 mmol) of PS-5 benzyl ester sulfoxide was used instead of
The same operation as in 11 was performed to obtain 8 mg of the target product (yield 39.5%). UVλ ^THF _nax nm (ε): 322 (8100). IRν ^CHCl3 _nax cm ^-1 : 1785 (β-lactam CO), 1720 (ester CO). _NMR ( _CDCl3 ) δ: 1.07 (3H, t, J=7.5Hz, _CH2CH3 ), 1.60-2.00 (2H, m _, CH2CH3 ₎ , 2.50-3.20 (3H, m, C-4 H, C-6
H), 3.88 (1H, dt, J=9Hz, J=3Hz, C-5
H), 5.36 (1H, d, J=12Hz, ArCH・H), 5.49 (1H, d, J=12Hz, ArCH・H ), 7.30~7.44 (2H, m, PyH), 8.50~8.70 (2H , m, PyH). Mass (m/e): 380 (M ⁺ ), 310 (M ⁺ −EtCH=C
=O). Example 14 3-(2-pyridylthio)-6-ethyl-7-oxo-1-azabicyclo[3.2.0]hepta-2
Preparation of -ene-2-carboxylic acid p-nitrobenzyl ester To a solution of 50 mg of PS-5-p-nitrobenzyl ester sulfoxide in 30 ml of dry DMF was added 17.8 mg of 2-pyridylmercaptan sodium salt under stirring at -45°. (produced from 14.8 mg of 2-mercaptopyridine and 6.4 mg of sodium hydride) in dried DMF1
ml solution was added dropwise. After stirring at the same temperature for 10 minutes, the reaction mixture was added to 100ml of benzene, 0.1M, PH6.8.
After washing four times with 100 ml of phosphate buffer, the solvent was dried over anhydrous sodium sulfate and evaporated under reduced pressure to obtain a yellow oil. This oil was subjected to column chromatography using silica gel (30 g, 2.6×
12cm) and benzene-acetone (40:1,
ν/ν), 14.9 mg of colorless powder was obtained. [α] ²² _D : +16.2℃ (C1.00, THF). IRν ^CHCl3 _nax cm ^-1 : 1780 (β-lactam CO), 1710 (ester CO). UVλ ^THE _nax nm (ε): 325 (16900), 269 (12100). NMR ( _CDCl3 ) δ: 1.00 (3H, t, J=7Hz _{, CH2CH3} ), 1.60-2.10 (2H, m _{, CH2CH3} ) 2.92 (1H, dd, J=18Hz, J=9Hz ,C-4
H), 3.22 (1H, dd, J=18Hz, J=9Hz, C-4
H), 3.07 (1H, dt, J=7Hz, J=3Hz, C-6
H), 3.92 (1H, dt, J=9Hz, J=3Hz, C-5
H), 5.26 (1H dd, J = 14Hz, ArC H H), 5.53 (1H, dd, J = 14Hz, ArCH H ), 7.40-7.80 (3H, m, PyH), 7.63 (2H, d, J = 8Hz, ArH), 8.20 (2H, d, J=8Hz, ArH), 8.58 (1H, dd, J=5Hz, J=2Hz, PyH). Mass (m/e): 425 (M ⁺ ), 355 (M ⁺ -EtCH=C=O). Example 15 3-(2-pyridylthio)-6-ethyl-7-oxo-1-azabicyclo[3.2.0]hepta-2
Production of -ene-2-carboxylic acid 3-(2-pyridylthio)-6-ethyl-7-oxo-1-azabicyclo[3.2.0]hepta-2-
Dissolve 70 ml of ene-2-carboxylic acid/p-nitrobenzyl ester in a mixed solvent of 3 ml of dioxane and 1.7 ml of 0.1MPH6.8 phosphate buffer, and dissolve 70 mg of platinum oxide.
and Pearl reduction equipment (hydrogen pressure 4.0Kgr/cm ² )
The mixture was subjected to a catalytic ring reaction for 4 hours at room temperature. After the reaction was completed, the catalyst was filtered off using a filter aid, and the filtrate was treated in the same manner as in Example 11 to obtain 19.9 mg of the purified target product (yield: 41.9%). UVλ _nax nm (ε): 305 (12900). NMR (CDCl ₃ ) δ: 0.99 (3H, t, J = 7.5Hz, CH ₂ CH ₃ ), 1.80 (2H, dq, J = 7.5Hz CH ₂ CH ₃ ), 2.82 (2H, d, J = 9.0Hz C-4 H), 3.26 (1H, dt, J=7.5Hz, J=3Hz, C-6
H), 3.98 (1H, dt, J=9.0Hz, J=3Hz, C-5
H) 7.41, 7.62, 7.83and8.45 (1H, each m,
ArH). Example 16 3-(2-pyrimidinylthio)-6-ethyl-7
-Preparation of oxo-1-azabicyclo[3.2.0]hept-2-ene-2-carboxylic acid p-nitrobenzyl ester Add 100 mg (0.223 mmol) of PS-5 p-nitrobenzyl sulfoxide to 15 ml of dry DMF. Dissolved and cooled to -35°C. While stirring, 33.5mg
A solution of 2-mercaptopyrimidine sodium salt (0.245 mmol) in 6 ml of dry DMF was added. After reacting for 40 minutes, pour the reaction solution into 80 ml of benzene and add 0.1 M phosphate buffer (PH8.4) (50 ml).
×3). After drying the organic layer over anhydrous sodium sulfate, the solvent was distilled off under reduced pressure to obtain a pale yellow oil. This was applied to a column (1.2 x 18 cm) filled with 10 g of silica gel and benzene.
It was developed with acetone (50:3v/v) solvent. Each fraction was searched by thin layer chromatography, fractions containing the target product were collected, and the solvent was concentrated under reduced pressure to obtain 30 mg of the target product (yield:
31.6%). [α] ²² _D : +38.4° (c0.47, THF). UVλ ^THF _nax nm (ε): 322 (9800), 268 (10300). IRν ^CHCl3 _nax cm ^-1 : 1780 (β-lactam CO), 1715 (ester CO). NMR ( _CDCl3 ) δ: 1.08 (3H, t, J=7Hz _{, CH2CH3} ), 1.68-2.00 (2H, m _{, CH2CH3} ), 3.00-4.10 (4H, m, C-4H ,C-5
H, C-6 H) 5.28 (1H, d, J=14Hz, ArC H・H), 5.52 (1H, d, J=14Hz, ArCH・H ), 7.04 (1H, t, J=6Hz, C- 5′ H), 7.62 (2H, d, J=9Hz, ArH), 8.20 (2H, d, J=9Hz, ArH), 8.56 (2H, d, J=6Hz, C-4 H, C-
6 H), Mass (m/e): 426 (M ⁺ ),
356 (M ⁺ -EtCH=C=O) Example 17 3-(2-pyrimidinylthio)-6-ethyl-7
-Production of oxo-1-azabicyclo[3.2.0]hept-2-ene-2-carboxylic acid 3-(2-pyrimidinylthio)-6-ethyl-7
-Oxo-1-azabicyclo[3.2.0]hepta-
Dissolve 63.4 mg of 2-ene-2-carboxylic acid/p-nitrobenzyl ester in 3.2 ml of dioxane and 1.2 ml of 0.1 M phosphate buffer (PH6.86), add 70 mg of platinum oxide, and use a Pearl reduction apparatus. (Hydrogen pressure 4
Kg/cm ² ) and reacted at room temperature for 4 hours. Separate the catalyst
The liquid was combined with the washing liquid, the pH was adjusted to 7.2, and the mixture was concentrated under reduced pressure to about two-thirds of its volume. After adding 2 g of sodium chloride to the concentrated solution, water was added to make up to 70 ml. Apply this solution to a Diamondion HP20AG column (1.1 x 20
cm), 0 to 50% (ν/ν) of the total volume of 300ml
Elution was performed by an acetone water gradient gradient method. Each fraction was searched using high-performance liquid chromatography, and fractions with ultraviolet absorption at λ _nax 300 nm were collected and freeze-dried to yield 16.8 mg.
The desired product was obtained (yield 39%). UVλ ^H2O* _nax nm (ε): 297 (6400). *0.1M phosphate buffer (PH7.0) NMR (D2O) δ: 1.02 (3H, t, J=7Hz, _{CH2CH3 ) 1.68-2.04} (2H, m _{, CH2CH3} ), 2.84-3.529 (3H, m, C-4 H, C-6
H) 4.14 (1H, dt, J=9Hz, J=8Hz, C-5
H), 7.35 (1H, t, J = 6Hz, C-5' H), 8.64 (2H, d, J = 6Hz, C-4' H, C-
6′ H). Example 18 3-(2-pyrimidinylthio)-6-ethyl-7
-Production of oxo-1-azabicyclo[3.2.0]-hept-2-ene-2-carboxylic acid benzyl ester 100 mg (0.223 mmole) of sulfoxide of PS-5 p-nitrobenzyl ester in Example 16
Example except that 44 mg (0.114 mmol) of PS-5 benzyl ester sulfoxide was used instead of
Purification was performed in the same manner as 16 to obtain 16 mg of the desired product. UVλ ^THF _nax nm (ε): 321 (9900). IRν ^CHCl3 _nax cm ^-1 : 1780 (β-lactam CO), 1715 (ester CO). NMR ( _CDCl3 ) δ: 1.04 (3H, t, J=7Hz _{, CH2CH3} ), 1.63-1.95 (2H, m _{, CH2CH3} ), 3.00-4.10 (4H, m, C-4H ,C-5
H, C-6 H), 5.34 (1H, d, J = 12Hz, ArC H H), 5.40 (1H, d, J = 12Hz, ArCH H ), 7.10-8.00 (8H, m, PyH, ArH ), 8.50-8.64 (1H, distorted d, PyH). Mass (m/e): 381 (M ⁺ ), 311 (M ⁺ −EtCH=C
=O). Example 19 Production of 6-ethyl-7-oxo-1-azabicyclo[3.2.0]hept-2-ene-2-carboxylic acid sodium salt 110 mg of PS-5 sodium salt was added to 0.01 M phosphate buffer (PH7. 3) After dissolving in 10 ml, 500 mg of palladium oxide was added and reduction was performed at room temperature and normal pressure (Ishii's reduction apparatus). After 6 hours of reaction, filter the reaction solution and apply 50 ml of the solution to a QAE Sephadex column (100
ml) and 300 ml of 0.01M phosphate buffer.
0.01M phosphate buffer containing 0.3M sodium chloride 300
Elution was performed using a 0-0.3M sodium chloride concentration gradient method using ml, and the first fraction of the target product having an absorption maximum at 268 nm was collected. 30ml of this fraction
Since there was a small amount of PS-5 in the, after adding 2g of sodium chloride,
Adsorb onto a column (20 ml), wash with 5 ml of 3% sodium chloride, then 20 ml of distilled water, and elute by concentration gradient method using water-3% acetone water to extract the fraction of the target substance with maximum absorption at 242 nm and 265 nm. was collected and freeze-dried to obtain 13.9 mg of the target product. UVλ ^H2O* _nax nm (ε): 265 (3000). *0.1M phosphate buffer (PH
7.3). NMR (D ₂ O) δ: 1.00 (3H, t, J = 7.5Hz, CH ₂ CH ₃ ), 1.80 (2H, dq, J = 7.5, J = 7.0Hz, CH ₂
CH ₃ ), 2.74 (1H, ddd, J=19.0Hz, J=9.5Hz, J=
2.5Hz, C-4 H・H), 2.94 (1H, ddd, J=19.0Hz, J=9.5Hz, J=
3.0Hz, C-4 H・H ), 3.27 (1H, dt, J=7.0Hz, J=3.0Hz, C-6
H), 4.05 (1H, dt, J=9.5Hz, J=3.0Hz, C-5
H), 6.24 (1H, t, J=3.0Hz, J=2.5Hz, C-3
H). Next, a method for producing a typical pharmaceutical preparation containing the compound of formula (-b) or a salt thereof of the present invention will be illustrated.
In addition, the compound number has the above-mentioned meaning. Example A: Capsule Ingredients Compound 1 per capsule 100 mg Lactose (Japanese Pharmacopoeia Law) Appropriate amount Magnesium stearate 1 mg The above antibiotic and excipients are ground in a mortar and mixed uniformly. Approximately 200 mg per capsule is packed into a No. 8 hard gelatin capsule with an enteric coating. Example B: Tablet Ingredients Compound 2 per tablet 200mg Lactose (Japanese Pharmacopoeia Law) 120mg Corn starch 175mg Magnesium stearate 5mg 500mg Thoroughly mix the antibiotic with half of the lactose and corn starch in the above mixing ratio. The mixture is granulated with 10% starch paste and sieved. Add the remainder of the corn starch and magnesium stearate to this, mix well, and make a 1cm diameter and weight
Form into 500mg tablets. This is coated with sugar and then coated with an enteric coating. Example C: Injection Ingredients 25 mg of compound 3 per vial 2 ml of physiological saline (Japanese Pharmacopoeia) Dissolve the above antibiotic in sterile water for injection.
Disinfect. The solution is dispensed into sterilized ampoules, water is removed aseptically by freeze-drying, and the opening of the ampule is sealed. Before use, open the package, add 2 ml of sterile physiological saline to the contents of the ampoule, and dissolve. Example D: Tablet Ingredients Compound 5 per tablet 20mg Cephalorizine 180mg Lactose (Japanese Pharmacopoeia) 120mg Corn starch 175mg Magnesium stearate 5mg 500mg Compound 5 and cephalorizine were mixed and the following formulation method was repeated in the same manner as in Example B. It is formulated, molded, sugar-coated and then coated with an enteric coating. Example E: Tablet Ingredients Compound 4 per tablet 10mg Aminobenzylpenicillin 190mg Lactose (Japanese Pharmacopoeia) 120mg Corn starch 175mg Magnesium stearate 5mg 500mg Compound 4 and aminobenzylpenicillin were mixed and the same method as in Example B was carried out. Formulated into a formulation. Example F: Capsule Ingredients Compound 5 per capsule 100 mg Lactose Adequate amount Magnesium stearate 1 mg The above antibiotic derivative and excipients are ground and mixed uniformly. Approximately 200 mg per capsule is packed into a No. 8 hard gelatin capsule with an enteric coating. Example G: Tablet Ingredients Compound 4 per tablet 200 mg Lactose (Japanese Pharmacopoeia) 120 mg Corn starch 175 mg Magnesium stearate 5 mg 500 mg The antibiotic derivative is thoroughly mixed with half of the lactose and corn starch in the above mixing ratio. mixture
Mix with 10% starch paste solution to granulate, and pass through a sieve. Add the remaining corn starch and magnesium stearate to this, mix well, and make a
cm, mold into tablets with a weight of 500 mg per tablet. This is coated with sugar and then coated with an enteric coating. Example H: Injection Ingredients Compound 1 per vial 25 mg 70% M-(β-hydroxyethyl)lactamide aqueous solution 2 ml The above antibiotic derivative is dissolved in sterile water for injection and sterilized by filtration. The solution is dispensed into sterilized ampoules, water is removed aseptically by freeze-drying, and the mouth of the ampule is sealed. Before use, open the ampoule, add 2 ml of a 70% aqueous solution of sterile N-(β-hydroxyethyl)lactamide to the contents of the ampoule, dissolve, and use. Example I: Tablet Ingredients Compound 3 per tablet 50mg Cephaloridine 150mg Lactose (Japanese Pharmacopoeia) 120mg Corn starch 175mg Magnesium stearate 5mg 500mg This antibiotic derivative and Cephaloridine were mixed,
Formulated and molded in the same manner as the formulation method of Example G,
After sugar coating, it is further coated with an enteric coating. Example J: Tablet Ingredients Compound 2 per tablet 20mg Aminobenzylpenicillin 180mg Lactose (Japanese Pharmacopoeia) 120mg Corn starch 175mg Magnesium stearate 5mg 500mg Compound 2 and aminobenzylpenicillin were mixed and prepared according to the method of [Tablet I]. Formulate in the same manner.

Claims (1)

[Claims] 1 The following formula () In the formula, R ₁ is a phenyl group, a nitrophenyl group,
a cyclohexyl group, a pyridyl group, or a pyrimidinyl group, and R ₂ represents a hydrogen atom, a benzyl group, or a nitrobenzyl group, and a salt of the compound of formula () when R ₂ is a hydrogen atom. 2 The compound of the formula () has the following formula (-a) The compound according to claim 1 and a salt thereof having a 5,6-trans configuration represented by: 3 The following formula (-b) In the formula, R ₁ is a phenyl group, a nitrophenyl group,
An antibacterial agent containing as an active ingredient a compound or a salt thereof representing a cyclohexyl group, a pyridyl group, or a pyrimidinyl group. 4 (A) The following formula () In the formula, R′ ₂ represents a benzyl group or a nitrobenzyl group, (i) A compound represented by the following formula () R ₁ −SH () In the formula, R ₁ is a phenyl group, a nitrophenyl group,
(B ) Then, the following formula (-d) is obtained: In the formula, R ₁ and R′ ₂ have the above-mentioned meanings, and the compound represented by the following formula is optionally subjected to hydrogenolysis. A method for producing a compound or a salt thereof, wherein R ₁ has the above-mentioned meaning, and R ₂ represents a hydrogen atom, a benzyl group, or a nitrobenzyl group.