JPS634540B2 - - Google Patents

Description

[Detailed description of the invention]

The present invention provides a novel β-
This invention relates to lactam compounds and their production methods. More specifically, the object of the present invention is of the formula () A compound or its salt represented by the formula () It can be produced by subjecting the compound represented by or a salt thereof to a ring reaction. Examples of the above reduction reaction include catalytic reduction reaction. This catalytic reduction reaction can be carried out by a conventional method, and examples of catalysts include platinum catalysts such as platinum wire, platinum sponge, platinum black, platinum oxide, and colloidal platinum, palladium sponge, palladium black, palladium oxide, palladium barium sulfate, and palladium Examples include palladium catalysts such as barium carbonate, palladium on carbon, palladium silica gel, and colloidal palladium, and nickel catalysts such as reduced nickel, nickel oxide, Raney nickel, and Soehara nickel. Examples of solvents that can be used include solvents that dissolve the raw material compound (), such as water or a mixture of water and a polar organic solvent such as dioxane, tetrahydrofuran, dimethylformamide, methanol, ethanol, and propanol. can be mentioned. This reaction is preferably carried out at 0 DEG to 50 DEG C. and under hydrogen pressure of 1 to 2 atmospheres. After the reaction is complete,
The object of the present invention () is obtained by removing the catalyst from the reaction mixture, for example, by filtration.
Unreacted raw materials can be easily removed by adsorbing them on a polystyrene adsorbent resin such as XAD-2 resin (Rohm & Haas, USA) and subjecting them to column chromatography using water or hydrous alcohol as the eluent. Can be separated from substances or reaction by-products. Typically, the fraction eluted from the XAD-2 resin is detected using a high performance liquid chromatography system using a 254 nm UV detector and a Microhobond Pack C ₁₈ (Waters Inc., USA). This is done by injecting a sample of Examples of the salts of compounds () and () include alkali metal salts such as sodium, potassium, and lithium, alkaline earth metal salts such as magnesium, calcium, and barium, and organic amine salts such as trimethylamine, triethylamine, and pyridine. The raw material compound () of the present invention is, for example, Streptomyces sp. C-19393 (FERM-P
No. 4774, IFO 13886, ATCC 31486) in a medium containing nutrients that can be used by microorganisms, and is called antibiotic C-19393S ₂ (Patent application 11560-1982). . The antibacterial spectra of the compounds obtained in the Examples are shown in Table 1.

[Table] Mu
(Salmonella typhimurium)

[Table] Note) Medium: Meat juice agar The compound () obtained by the present invention exhibits antibacterial activity against Gram-positive bacteria and Gram-negative bacteria, as is clear from the above antibacterial spectrum. Therefore, the compounds () can be used to treat bacterial infections in mammals (eg, mice, rats, dogs, humans) and domestic animals (eg, chickens, ducks). In order to use the compound () as a therapeutic agent for E. coli infection, for example, the compound () can be dissolved in physiological saline and administered parenterally subcutaneously or intramuscularly at 0.1 to 200 mg/Kg/day, preferably as an injection. is administered at a dose of 1 to 50 mg/Kg/day. Also, as an oral agent,
Compound () is mixed with lactose to make capsules,
The compound () is administered at 1 to 500 mg/Kg/day, preferably 5 to 200 mg/Kg/day. Moreover, the compound () obtained by the present invention
can be used as a fungicide. For example, a solution prepared by dissolving the compound () in distilled water at a concentration of 0.01 to 1.0 W/V%, or a solution containing 0.5 to 50 mg of the compound () per 1 g based on petrolatum or lanolin,
Preferably, the ointment containing 2 to 20 mg can be used for sterilizing and disinfecting the hands, feet, eyes, ears, etc. of the above-mentioned animals. Compound () exhibits a beta-lactamase inhibitory effect and significantly enhances the susceptibility of penicillin- or cephalosporin-resistant bacteria to ampicillin and cefotiam due to the ability to produce beta-lactamase. Therefore, by combining the compound () with penicillins or cephalosporins, it is possible to treat bacteria in mammals (e.g. mice, rats, dogs, humans) and poultry (e.g. chickens, ducks), especially beta-lactam antibiotics. It can be used to treat infections caused by resistant bacteria. Compound () can be used in combination with other beta-lactam drugs, for example, to treat infections caused by beta-lactam antibiotic-resistant E. coli.
For example, the same amount of compound (2) and ampicillin are dissolved in physiological saline and administered as an injection parenterally subcutaneously or intramuscularly at 0.1 to 20 mg/Kg/day, preferably 0.5 to 5 mg/Kg/day. In addition, as an oral preparation, a capsule containing the same amount of the compound (2) and cephalexin is administered at 1 to 200 mg/Kg/day, preferably 5 to 100 mg/Kg/day. As a bactericidal agent, for example, compound ()0.1~
10W/V% concentration and benzylpenicillin 0.1~
Hands and feet of the above-mentioned animals, either as a solution in an aqueous solution containing a concentration of 1.0 W/V% or as an ointment based on petrolatum or lanolin and containing 5 to 20 mg of the compound () and 5 to 20 mg of benzylpenicillin per gram. It can be used to sterilize and disinfect the eyes, ears, etc. Compound () is also an extremely promising compound as a synthetic intermediate for new pharmaceuticals. Aqueous solutions of the compounds of the present invention are stable in the neutral pH range. Next, the present invention will be explained in more detail using reference examples and examples, but the present invention is not limited thereto. Percentages in reference examples indicate weight/volume % unless otherwise specified. Reference example Streptomyces sp. C-19393
(FERM-P No.4774, IFO 13886, ATCC
31486) into a 1 volume Erlenmeyer flask.
Grow on T medium* and allow spores to settle. The spores are then suspended in sterile water to a viable cell count of 1.2 x 10 ⁸ /ml. The spore suspension was diluted 10 times with sterile water, 1 ml of it was inoculated into 40 ml of seed medium in a 200 ml Erlenmeyer flask, and cultured on a rotary shaking culture machine at 28°C for 2 days. Inoculate 500 ml of seed medium in a Sakaguchi shake flask, culture on a reciprocating shaker for 2 days at 28°C, and transfer the culture to 15 ml of Actocol in a 50-volume stainless steel fermenter.
Transplant into seed medium containing 30% and aerate at 28°C with 30% aeration.
The cells were cultured for 3 days with stirring at 280 rpm. The culture solution was then transferred to a 1.2 m ³ main medium in a 2 m ³ fermentor, and cultured for 5 days at 30° C. with an aeration rate of 840/min and stirring at 180 rpm. The seed medium has a pH of 7.0 and consists of 20g of glucose, 30g of soluble starch, 2% oatmeal, 2% tomato paste, 0.2% Bovril (manufactured by Bovril in the UK), and 2% agar.
medium, g, raw soybean flour 10g, corn steep,
10 g of Ricart, 5 g of polypeptone (manufactured by Daigo Nutritional Chemical Co., Ltd.), 3 g of salt, 5 g of precipitated calcium carbonate
The pH was adjusted to 7.0 before sterilization. The main medium contains 30g glucose and soluble starch per serving.
30g, defatted soybean flour 15g, cotton seed flour 15g, potassium dihydrogen phosphate 0.25g, potassium monohydrogen phosphate 0.6g,
It contained 0.002 g of cobalt chloride and 0.5 g of Actocol, and its pH was adjusted to 7.0 before sterilization. All media were steam sterilized at 120°C for 20 minutes. Hyflo Super Cell was added to the culture solution obtained as described above and filtered to obtain solution 1230.
After adjusting the pH of the liquid to 6.3, it was passed through a column packed with 100% activated carbon. Antibiotics were eluted with 300 ml of water. Dowex the eluate 1 x 2 (C1 ^- type, 2)
After washing with 6 portions of water, the column was eluted with 32 portions of 5% saline. After adjusting the eluate to pH 5, it was passed through a column packed with activated carbon 4. water 12
After washing with 8% isobutanol water 7 and 8%
The antibiotic was eluted with isobutanol-N/20 ammonia water 12 times. After concentrating the eluate to 150 ml under reduced pressure, 1350 ml of methanol was added to remove the resulting precipitate. After concentrating the liquid to 200ml, add 300ml of DEAE A25 (CI ^- type)
0.1M and 0.2M
-After washing the column with saline (900ml each), 0.4M-
Antibiotics were eluted with 1500 ml of saline. PH the eluate
After adjusting the concentration to 500 ml, the column was passed through a column packed with 500 ml of activated carbon, washed with 1.5 ml of water, and eluted with 2.5 ml of 8% iso-butanol-N/20 aqueous ammonia. The eluate was concentrated to dryness and acetone was added to obtain 2.4 g of slightly yellow powder. Dissolve the obtained powder in a small amount of water and add Amberlite XAD-(100-200
Pass through a column packed with 1.2
It was eluted and fractionated with water. Collect and concentrate the antibacterial fraction, and use the concentrated liquid as QAE-Sephadex A25 (CI ^- type)
Pass through a column packed with 200ml, wash the column with 600ml of 0.1M saline, and then add 1.2ml of 0.2M saline.
It was eluted. After adjusting the eluate to pH 5, add activated carbon 600
After passing through a column packed with 1.8 ml of water and washing with 1.8 ml of water, the solution was eluted with 3 ml of 8% iso-butanol-N/20 ammonia water. The eluate was concentrated to dryness and acetone was added to obtain 1.07 g of powder. 620 mg of the obtained powder was dissolved in a small amount of water, passed through a column packed with 360 ml of Amberlite XAD-(100-200 mesh), eluted with water, and fractionated. Each of the fractions that were found to have antibacterial properties was subjected to the liquid chromatography analysis described above, and the portions showing a single peak were collected, concentrated, dried, and acetone added.
Antibiotic C-19393S ₂ disodium salt white powder
Obtained 136 mg. The specific rotation of this powder is [α] ²² _D −
The temperature was 152°±15° (C=0.5, water). Example Antibiotic C-19393S ₂ Disodium salt crude powder (30% purity, 60mg) was added to 10% methanol water (20ml)
This solution was added to a mixture of 10% methanol water (10 ml) and 10% palladium-carbon (20 mg) that had been hydrogenated for 30 minutes. The mixture was then reduced with hydrogen at 1 atm for 3 hours at room temperature, the catalyst was filtered off, and the filtrate was concentrated in vacuo to a volume of 2 ml. This solution was passed through a column (50 ml) of XAD-2 (100 to 200 meshes) to adsorb the target compound (), and then eluted with water. 45ml
Collect the elution fractions containing the target compound () up to 150 ml and freeze-dry [5R, 6R]-3-
[(E)-2-acetamidoethenylthio]-6-
7.3 mg of powder of disodium [1-(hydroxysulfonyloxy)-1-methylethyl]-7-oxo-1-azabicyclo[3,2,0]hept-2-ene-2-carboxylate was obtained.・UV: λmax (H ₂ O) 228 and 309 nm ・IR: ν: max (KBr) 1760, 1620, 1240, 1050
cm ^-1 Thin layer chromatography [cellulose f (manufactured by Tokyo Kasei Co., Ltd.)]: R _f = 0.65 (solvent system: propanol: water = 4:1) - High performance liquid chromatography (manufactured by Waters Corporation): Rt = 4.4 minutes [Micro Bonder Pack C18/
14% methanol-0.02M-phosphate buffer (PH
6.3), 2 ml/min/cm (200 PSi)], but under the same conditions, the Rt of the raw material compound () was 2.2 minutes. NMR; δ (100MHZ, D ₂ O, TMs): 1.63 (3H,
S, _C8 - _CH3 ), 1.70 (3H, S, _C8 - _CH3 ), 2.10
(3H, S, COCH ₃ ), 3.05 (1H, dd, j=10,
19, _C4 - H ), 3.82 (1H, dd, 10.5, 19 _C4 - H ),
3.88 (1H, d, J = 6, C ₆ - H ), 4.20 (1H, m,
_C5 - H ), 6.10 (1H, d, J=14, N- CH =),
7.20 (1H, d, S- CH =)

Claims (1)

[Claims] 1 formula Compounds represented by and their salts. (2) Equation A formula characterized by subjecting a compound represented by or a salt thereof to a reduction reaction A method for producing a compound represented by or a salt thereof.