JP2013095719A - Antibacterial agent (against bacteria of genus clostridium) and manufacturing method for the same - Google Patents

Abstract

PROBLEM TO BE SOLVED: To provide an antibacterial agent against bacteria of the genus Clostridium.SOLUTION: Compounds represented by 3-acyl-4-hydroxy-6-methylpyrones, 3-acyltetronic acids and/or 3-acyl-4-hydroxycoumarins are used for an antibacterial agent against bacteria of the genus Clostridium. These compounds are particularly selective for Clostridium difficile and/or Clostridium perfringens among bacteria of the genus clostridium.

Description

  The present invention relates to an antibacterial agent, a medicine, a treatment method, a method for use as an antibacterial agent, and a method for producing an antibacterial agent, and in particular, an antibacterial agent, medicine, and treatment for Clostridium bacteria that are selective for Clostridium bacteria The present invention relates to a method, a method for use as an antibacterial agent, and a method for producing an antibacterial agent.

In recent years, infections caused by bacteria belonging to the genus Clostridium, which are pathogenic enteric bacteria, have become a problem.
Bacteria belonging to the genus Clostridium are positive by Gram staining, and are characterized by being anaerobic and forming spores. In addition, some Clostridium bacteria exist as resident bacteria in the intestines of humans and animals, and pathogenic bacteria also exist. Among these, infectious diseases such as colitis caused by Clostridium difficile (hereinafter referred to as C. difficile) or Clostridium perfringens are particularly problematic.

Here, more than 400 types of intestinal bacteria in the intestine of a healthy person are well-balanced to form an “intestinal flora (flower field)”. In an environment where this “intestinal flora” is functioning normally, C.I. Proliferation is suppressed even if difficile is infected. However, when an antibacterial agent having a broad antibacterial spectrum is taken for the treatment of other diseases, intestinal bacteria that are useful for digestion and absorption are also sterilized, and the balance of the intestinal flora is lost. In addition, C. and C. When difficile grows abnormally, it produces toxins inside the large intestine that can cause severe diarrhea as well as death.
In many developed countries such as the United States, Canada and Europe, C.I. Nosocomial infections due to difficile have become a serious problem. For example, in the United States alone, approximately 700,000 cases occur annually, and the cost of the insurance medical system amounts to $ 3.2 billion per year. In the EU member states, the associated medical costs are estimated at $ 4.4 billion per year.
This C.I. Most antibacterial agents are not effective against difficile infections, and vancomycin and metronidazole are slightly used as therapeutic agents.
Vancomycin is a glycopeptide antibiotic having a bacterial cell wall synthase inhibitory activity and is characterized by high antibacterial activity in a wide antibacterial spectrum. Metronidazole is an antibacterial drug approved as a secondary sterilization therapy against Helicobacter pylori. It is also used against difficile infections.
However, C.I. Difficile infection is characterized by high morbidity and mortality.
In Japan, C.I. Although difficile is not tested for infection, it is thought that there are many potential infected persons.

  Clostridium perfringens is also a kind of resident bacteria that inhabit the intestines of humans and animals. Clostridium perfringens also produces toxins, especially during spore formation. For this reason, when reheating of the foodstuff containing meat is insufficient, it may become a cause of large-scale food poisoning.

Here, the conventional C.I. As an antibacterial agent for difficile, referring to Patent Document 1, a hop extract or a hop extract component of 1 ppm or more is carried into the stomach or intestine to obtain C.I. A technique characterized by inhibiting the growth of difficile has been disclosed (hereinafter referred to as Conventional Technique 1).
According to the hop extract of prior art 1, Clostridium botulinum and C.I. An antibacterial agent that inhibits the growth of difficile can be provided.

  Further, referring to Non-Patent Documents 1 and 2, known lactones exhibiting antibacterial activity are described.

Japanese Patent Laid-Open No. 11-222104

Tadao Toda, Toru Tokunaga, Tsutomu Ouchida, Kenichi Shiroya, Takao Nakamoto, “In vitro antibacterial action of pyrone derivatives (Studies on chemotherapeutic agents)”, CHEMOTHERAPY, Japan, MAR. 1958, Vol. 6, no. 2, P.I. 91-95 Mikio Namiki, Masao Nomoto, Kazuko Yamamoto, Ryota Shimose, Synthesis of Hydroacyl Succinic Acid and Carboxylic Acid (Part 4) Relationship between Side Chain Elongation and Antibacterial Activity " , Japan, Japan Agricultural Chemical Society Tokyo Branch Regular Meeting, Agricultural Chemistry Magazine, July 1951, Volume 26, 211-218

However, the antibacterial agent of Prior Art 1 is C.I. For difficile infection, pharmacological activity was low for use as an effective therapeutic agent.
In addition, C.I. For difficile infections, most antibacterial agents such as antibiotics did not show effectiveness, and vancomycin and metronidazole were only used. However, recurrence and drug resistance when taking the medicine have been serious problems.
For this reason, a new C.I. There has been a long-awaited development of antibacterial agents that can be used to treat difficile infections. Non-patent documents 1 and 2 do not describe a bacterium belonging to the genus Clostridium.

  This invention is made | formed in view of such a condition, and makes it a subject to eliminate the above-mentioned subject.

で表される（ただし、Ｒは、炭素数４〜３０の直鎖状又は分岐鎖状又は環を形成していてもよいアルキル基である）ことを特徴とする。
本発明の抗菌剤は、前記３−アシルテトロン酸類は、式

で表される（ただし、Ｒは、炭素数４〜３０の直鎖状又は分岐鎖状又は環を形成していてもよいアルキル基である）ことを特徴とする。
本発明の抗菌剤は、前記３−アシル−４−ヒドロキシクマリン類は、式

で表される（ただし、Ｒは、炭素数４〜３０の直鎖状又は分岐鎖状又は環を形成していてもよいアルキル基である）ことを特徴とする。
本発明の医薬は、前記抗菌剤を用いたことを特徴とするクロストリジウム属細菌用の大腸炎治療用の医薬であることを特徴とする。
本発明の非ヒト動物の大腸炎の治療方法は、３−アシル−４−ヒドロキシ−６−メチルピロン類で表される化合物、３−アシルテトロン酸類で表される化合物、及び／又は３−アシル−４−ヒドロキシクマリン類で表される化合物を投与することを特徴とする。
本発明の抗菌剤として用いる方法は、３−アシル−４−ヒドロキシ−６−メチルピロン類で表される化合物、３−アシルテトロン酸類で表される化合物、及び／又は３−アシル−４−ヒドロキシクマリン類で表される化合物をクロストリジウム属（Ｃｌｏｓｔｒｉｄｉｕｍ）細菌用の抗菌剤として用いる使用方法であることを特徴とする。
本発明の抗菌剤の製造方法は、４-ヒドロキシ-６-メチルピロン、テトロン酸、及び／又は４-ヒドロキシクマリンの１種を基質とし、有機溶媒中、アミン塩基存在下において、酸クロリド又は酸無水物と前記基質とを反応させ、０〜５０℃で１２〜２４時間撹拌し、３−アシル−４−ヒドロキシ−６−メチルピロン類、３−アシルテトロン酸類、及び／又は３−アシル−４−ヒドロキシクマリン類で表される化合物を合成するクロストリジウム属（Ｃｌｏｓｔｒｉｄｉｕｍ）細菌用の抗菌剤の製造方法であることを特徴とする。
本発明の抗菌剤の製造方法は、４-ヒドロキシ-６-メチルピロン、テトロン酸、及び／又は４-ヒドロキシクマリンを基質とし、有機溶媒中、縮合剤およびアミン塩基存在下において、カルボン酸と前記基質とを反応させ、０〜５０℃で１２〜２４時間撹拌し、３−アシル−４−ヒドロキシ−６−メチルピロン類、３−アシルテトロン酸類、及び３−アシル−４−ヒドロキシクマリン類で表される化合物を合成するクロストリジウム属（Ｃｌｏｓｔｒｉｄｉｕｍ）細菌用の抗菌剤の製造方法であることを特徴とする。 The antibacterial agent of the present invention is represented by a compound represented by 3-acyl-4-hydroxy-6-methylpyrones, a compound represented by 3-acyltetronic acids, and / or 3-acyl-4-hydroxycoumarins. It is characterized by being an antibacterial agent for Clostridium bacteria containing the compound.
In the antibacterial agent of the present invention, the genus Clostridium is Clostridium difficile and / or Clostridium perfringens.
In the antibacterial agent of the present invention, the 3-acyl-4-hydroxy-6-methylpyrone is represented by the formula:

(Wherein R is a linear or branched chain or alkyl group which may form a ring having 4 to 30 carbon atoms).
In the antibacterial agent of the present invention, the 3-acyltetronic acids have the formula

(Wherein R is a linear or branched chain or alkyl group which may form a ring having 4 to 30 carbon atoms).
In the antibacterial agent of the present invention, the 3-acyl-4-hydroxycoumarins have the formula

(Wherein R is a linear or branched chain or alkyl group which may form a ring having 4 to 30 carbon atoms).
The medicament of the present invention is a medicament for treating colitis for Clostridium bacteria characterized by using the antibacterial agent.
The method for treating colitis in a non-human animal of the present invention comprises a compound represented by 3-acyl-4-hydroxy-6-methylpyrones, a compound represented by 3-acyltetronic acids, and / or 3-acyl- A compound represented by 4-hydroxycoumarins is administered.
The method used as the antibacterial agent of the present invention includes a compound represented by 3-acyl-4-hydroxy-6-methylpyrones, a compound represented by 3-acyltetronic acids, and / or 3-acyl-4-hydroxycoumarin. It is characterized by being a method of using a compound represented by the class as an antibacterial agent for Clostridium bacteria.
The method for producing an antibacterial agent of the present invention uses 4-hydroxy-6-methylpyrone, tetronic acid, and / or 4-hydroxycoumarin as a substrate, and acid chloride or acid anhydride in an organic solvent in the presence of an amine base. The product is reacted with the substrate and stirred at 0 to 50 ° C. for 12 to 24 hours to give 3-acyl-4-hydroxy-6-methylpyrones, 3-acyltetronic acids, and / or 3-acyl-4-hydroxy It is a method for producing an antibacterial agent for Clostridium bacteria that synthesizes compounds represented by coumarins.
The method for producing an antibacterial agent of the present invention uses 4-hydroxy-6-methylpyrone, tetronic acid, and / or 4-hydroxycoumarin as a substrate, and the carboxylic acid and the substrate in an organic solvent in the presence of a condensing agent and an amine base. And is stirred at 0 to 50 ° C. for 12 to 24 hours, and is represented by 3-acyl-4-hydroxy-6-methylpyrones, 3-acyltetronic acids, and 3-acyl-4-hydroxycoumarins It is a method for producing an antibacterial agent for Clostridium bacteria which synthesizes a compound.

  According to the present invention, a bacterium belonging to the genus Clostridium that is highly active against a bacterium belonging to the genus Clostridium, has a relatively low activity against other enteric bacteria, and has a selectivity that hardly affects the intestinal flora. An antibacterial agent can be provided.

<Embodiment>
As mentioned above, Clostridium bacteria, particularly C.I. There are very few antibacterial compounds that are selective against difficile and Clostridium perfringens and do not disrupt the balance of the intestinal flora, and there have been no examples of approval in Japan.
For this reason, the inventors of the present invention conducted intensive experiments and research to evaluate antibacterial activity against bacteria of the genus Clostridium. As a result, some known lactones have been found to be pathogenic enterobacteria C. The present invention was completed by finding that it has high antibacterial activity against difficile, Clostridium perfringens and the like, and antibacterial selectivity in enteric bacteria. These known lactones are several types of compounds described in Non-Patent Documents 1 and 2.
That is, the lactone compound having antibacterial activity selective to the bacterium belonging to the genus Clostridium according to the embodiment of the present invention is a 3-acyl-4-hydroxy-6-methylpyrone (3-Acyl-4-hydroxy-6-methylpyrone). ), 3-acyltetronic acids (3-Acyltetranic acid), and / or 3-acyl-4-hydroxycoumarins (3-Acyl-4-hydroxycoumarin).
That is, these compounds are C.I. In the in vitro antibacterial activity evaluation against difficile, it was equivalent to or higher in activity than vancomycin, which is a general-purpose drug.
Specifically, these compounds have the effect of selecting bacteria of the genus Clostridium at about 0.5 μg / mL to 64 μg / mL.

  Here, as the 3-acyl-4-hydroxy-6-methylpyrones of the antibacterial compound according to the embodiment of the present invention, a compound represented by the following general formula (1) can be used.

（ただし、Ｒは、炭素数４〜３０の直鎖状又は分岐鎖状又は環を形成していてもよいアルキル基である）。

(However, R is an alkyl group which may form a linear or branched chain or ring having 4 to 30 carbon atoms).

（ただし、Ｒは、炭素数４〜３０の直鎖状又は分岐鎖状又は環を形成していてもよいアルキル基である）。 In addition, as the 3-acyltetronic acids of the antibacterial compound according to the embodiment of the present invention, a compound represented by the following general formula (2) can be used.

(However, R is an alkyl group which may form a linear or branched chain or ring having 4 to 30 carbon atoms).

  Moreover, the compound represented by the following general formula (3) can be used for the 3-acyl-4-hydroxycoumarins of the antibacterial compound according to the embodiment of the present invention.

（ただし、Ｒは、炭素数４〜３０の直鎖状又は分岐鎖状又は環を形成していてもよいアルキル基である）。

(However, R is an alkyl group which may form a linear or branched chain or ring having 4 to 30 carbon atoms).

Thus, R which is a substituent in the above general formulas (1) to (3) uses an alkyl group having 4 to 30 carbon atoms, which may form a linear or branched chain or a ring.
The reason why an alkyl group which may form a linear or branched chain or ring having 4 to 30 carbon atoms can be used for R as the substituent is 3-acyl-4-hydroxy having 1 carbon. It can be mentioned that the present inventors have found that -6-methylpyrones, 3-acyltetronic acids, and 3-acyl-4-hydroxycoumarins exhibit an antibacterial action selectively against bacteria of the genus Clostridium. On top of this, the alkyl group which may form a linear or branched chain or ring having 4 to 30 carbon atoms reduces the polarity and greatly enhances the action of inhibiting the metabolism of the cell wall of Clostridium bacteria. It is thought to do. When the number of carbon atoms is reduced to less than 4, it is considered that the fat solubility is reduced and it is difficult to be taken into bacteria, resulting in a decrease in antibacterial properties. Conversely, if the number of carbons is too large, the fat solubility will increase and there will be an effect on the melting point, solubility, etc., so it will be less likely to be taken into bacteria and the antibacterial properties will be reduced.
Experimentally, the carbon number of the alkyl group which may form a linear or branched chain or ring is more preferably 4 to 20 carbon atoms, and more preferably 5 to 18 carbon atoms for the above reasons. I know. In addition, it has 9 to 18 carbon atoms and belongs to the genus Clostridium, particularly C. The effect of the highest antibacterial activity and selectivity against difficile and Clostridium perfringens is obtained. For this reason, in the following Example, the example whose carbon number is 9 and 13 was shown.
That is, the compounds according to the above general formulas (1) to (3) according to the embodiment of the present invention are highly active against Clostridium bacteria as compared to vancomycin which is a general-purpose therapeutic agent. In addition to this, it can be used as an antibacterial agent and a therapeutic agent that have a relatively low activity against other intestinal bacteria and have a selectivity that hardly affects the intestinal flora.

  Here, the antibacterial agent according to the embodiment of the present invention is an arbitrary pharmaceutically acceptable carrier (for example, physiological saline, isotonic solution containing an adjuvant, such as D-sorbitol, D-mannose, D-mannitol). Suitable solubilizers such as alcohols, specifically ethanol, polyalcohols such as propylene glycol, polyethylene glycol, nonionic surfactants such as polysorbate 80 (TM), HCO-50. And the like, but is not limited thereto). Moreover, a suitable excipient | filler etc. may be included.

In addition, the antibacterial agent according to the embodiment of the present invention may contain an appropriate pharmaceutically acceptable carrier in order to prepare a pharmaceutically acceptable carrier.
This carrier may include biocompatible materials such as silicone, collagen, and gelatin. Alternatively, various emulsions may be used.
Further, for example, one or two or more pharmaceutical additives selected from diluents, fragrances, preservatives, excipients, disintegrants, lubricants, binders, emulsifiers, plasticizers and the like may be contained. Good.

In addition, the antibacterial agent according to the embodiment of the present invention can be formulated using a pharmaceutically acceptable carrier well known in the art in a dosage form suitable for oral administration.
Although the administration route of the pharmaceutical composition according to the embodiment of the present invention is not particularly limited, it can be administered parenterally.
Parenteral administration includes, for example, intravenous, intraarterial, subcutaneous, intradermal, intramuscular or intraperitoneal administration.

Further, the antibacterial agent according to the embodiment of the present invention can be used for the treatment of severe colitis such as pseudomembranous colitis.
In addition, the antibacterial agent according to the embodiment of the present invention can be used for treatment of cases after antibiotic treatment and various cases including nosocomial infections for patients whose immunity is reduced by various treatments including anticancer agents.

  In addition, in order to use the antibacterial agent according to the embodiment of the present invention for the above-mentioned treatment, the administration interval and the dosage should be appropriately selected and changed according to various conditions such as the state of the disease and the condition of the subject. Is possible.

In addition, the dose and frequency of administration of the antibacterial agent according to the embodiment of the present invention depend on the purpose of administration, and further according to various conditions such as the age and weight of the patient, symptoms, and severity of the disease. It is possible to select and change as appropriate.
The number of administrations and the period are monitored for the administration state, and repeated or repeated administration depending on the state.

  Moreover, the antibacterial agent which concerns on embodiment of this invention can also be used together with another composition etc. In addition, the composition of the present invention may be administered simultaneously with other compositions, or may be administered at intervals, but the administration order is not particularly limited.

  In the embodiment of the present invention, the period during which the disease is improved or alleviated is not particularly limited. However, it may be temporary improvement or reduction, or may be improvement or reduction for a certain period.

In addition, the antibacterial agent according to the embodiment of the present invention is not particularly limited to humans, and includes, for example, livestock animal species and wild animals.
Therefore, the antibacterial agent according to the embodiment of the present invention can be widely used for animal treatment, livestock growth enhancement, and the like.
Moreover, it can also be included in foods such as health foods, animal feeds, or diets for disease prevention and health promotion.

Moreover, the antibacterial agent which concerns on embodiment of this invention can be added to a foodstuff, feed, and various products according to a conventional method. The addition amount at this time can be appropriately selected according to the type of product. However, when the concentration per mass% of the product is about 0.5 μg / mL to 64 μg / mL, the effect of Clostridium spp. It is done.
Moreover, the antibacterial agent which concerns on embodiment of this invention can be utilized with a various form. For example, it can be blended in foods, cosmetics, soft drinks, Japanese confectionery, Western confectionery, soft drinks, gargles, hand-washing disinfectants, wet tissues, toiletry products, disinfectants and the like.
In addition, as an application product of the antibacterial agent according to the embodiment of the present invention, specifically, food packaging materials, food storage containers, pickles, livestock meat products, fish meat products and other foods, livestock feed, pet food, etc. Can be used.

Moreover, it is possible to mix | blend an additive with an antibacterial agent which concerns on embodiment of this invention suitably according to a conventional method unless it is unsuitable.
The additives are not necessarily limited to one, but may be used in combination, such as product type, composition, expected contamination or spoilage microorganism, pH, water activity, required storage temperature. They can be used in combination depending on the storage period.

  In addition, the antibacterial agent according to the embodiment of the present invention is stable against heat, acid, and the like, reduces the number of living bacteria by heating food, and further contains the extract of the present invention. By using the agent, the storage stability can be effectively enhanced. In particular, antibacterial activity can be effectively obtained even against bacteria of the genus Clostridium that survive by spore formation even under high temperature sterilization.

[Method for Producing Lactone Compound of Antibacterial Agent According to Embodiment of the Present Invention]
3-Acyl-4-hydroxy-6-methylpyrones, 3-acyltetronic acids, and / or 3-acyls, which are lactone compounds having antibacterial activity selective to Clostridium bacteria according to embodiments of the present invention -4-Hydroxycoumarins can be synthesized and produced by the following synthesis method (A) or synthesis method (B) using commercially available compounds.
By these synthesis methods, it is possible to obtain a high-quality compound having a higher purity and a higher yield than those of Non-Patent Documents 1 and 2.

Synthesis method (A):
The substrate is synthesized by reacting an acid chloride or acid anhydride in an organic solvent in the presence of an amine base and stirring the substrate at a temperature range of 0 to 50 ° C. for 12 to 24 hours.
Examples of the organic solvent to be used include alkane chlorides such as dichloromethane and dichloroethane, nitriles such as acetonitrile and propionitrile, esters such as ethyl acetate, and ethers such as tetrahydrofuran and dioxane, preferably dichloromethane and acetonitrile are used. .
Examples of the amine base to be used include tertiary amines such as triethylamine and diisopropylethylamine, pyridines such as pyridine and 4- (dimethylamino) pyridine, and 4- (dimethylamino) pyridine is more preferably used.

Synthesis method (B):
The substrate is synthesized by allowing a carboxylic acid to act in an organic solvent in the presence of a condensing agent and an amine base, and stirring for 12 to 24 hours in a temperature range of 0 to 50 ° C.
Examples of the organic solvent to be used include alkane chlorides such as dichloromethane and dichloroethane, nitriles such as acetonitrile and propionitrile, esters such as ethyl acetate, and ethers such as tetrahydrofuran and dioxane, preferably dichloromethane and acetonitrile are used. .
As the condensing agent to be used, substituted carbodiimides such as N, N′-dicyclohexylcarbodiimide can be used, and more preferably N- [3- (dimethylamino)] propyl-N′-ethylcarbodiimide and its hydrochloric acid. Use salt.
Examples of the amine base to be used include tertiary amines such as triethylamine and diisopropylethylamine, pyridines such as pyridine and 4- (dimethylamino) pyridine, and 4- (dimethylamino) pyridine is more preferably used.

In each synthesis method (A) and (B):
3-Acyl-4-hydroxy-6-methylpyrones can be synthesized using 4-hydroxy-6-methylpyrone as a substrate.
Further, 3-acyltetronic acids can be synthesized using tetronic acid as a substrate.
Further, 3-acyl-4-hydroxycoumarins can be synthesized using 4-hydroxycoumarin as a substrate.

  EXAMPLES Next, the present invention will be further described with reference to examples, but the following specific examples are not intended to limit the present invention.

[Example 1]
As 3-acyl-4-hydroxy-6-methylpyrones, 3-decanoyl-4-hydroxy-6-methylpyrone of the following formula (I) was synthesized and used.
This 3-decanoyl-4-hydroxy-6-methylpyrone is an example having a linear alkyl group having 9 carbon atoms as R as a substituent in the general formula (1) of the above embodiment.

(Method for producing the compound of Example 1 in the synthesis method (A))
4-Hydroxy-6-methylpyrone (100 mg, 0.79 mmol,), 4- (dimethylamino) pyridine (145 mg, 1.18 mmol 1.5 equiv.) Was dissolved in dichloromethane (4 mL) and decanoic anhydride (284 mg, 0.87 mmol, 1.1 equiv.) Was added at room temperature.
After stirring at room temperature for 12 hours, 5% aqueous citric acid solution (10 mL) was added, and the mixture was extracted with ethyl acetate (20 mL).
The resulting organic layer was separated and dried over anhydrous sodium sulfate. The solution was concentrated and purified by silica gel column chromatography using an ethyl acetate-hexane mixed solvent to give 3-decanoyl-4-hydroxy-6-methylpyrone (137 mg, 62%) as a white solid.
In addition, as for each substrate and reagent, commercially available products such as those manufactured by Wako Pure Chemical Industries, Ltd. were used. The same applies hereinafter.

(Method for producing the compound of Example 1 in the synthesis method (B))
4-hydroxy-6-methylpyrone (100 mg, 0.79 mmol), decanoic acid (150 mg, 0.87 mmol, 1.1 equiv.), 4- (dimethylamino) pyridine (145 mg, 1.18 mmol 1.5 equiv.). Dissolved in dichloromethane (4 mL), N- [3- (dimethylamino)] propyl-N′-ethylcarbodiimide (226 mg, 1.18 mmol 1.5 equiv.) Was added at room temperature.
After stirring at room temperature for 12 hours, 5% aqueous citric acid solution (10 mL) was added, and the mixture was extracted with ethyl acetate (20 mL).
The resulting organic layer was separated and dried over anhydrous sodium sulfate. The solution was concentrated and purified by silica gel column chromatography using an ethyl acetate-hexane mixed solvent to give 3-decanoyl-4-hydroxy-6-methylpyrone (170 mg, 77%) as a white solid.

Regarding 3-decanoyl-4-hydroxy-6-methylpyrone according to the synthesis method (A) or (B) of Example 1, the following results were obtained by commercially available NMR.
¹ H NMR (300 MHz, CDCl 3) δ = 0.88 (t, 3H, J = 6.9 Hz), 1.20-1.42 (m, 14H), 1.58-1.80 (m, 2H) ), 2.54 (s, 3H), 2.82 (t, 2H, J = 7.8 Hz), 5.71 (s, 1H)

[Example 2]
As 3-acyltetronic acids, 3-decanoyltetronic acid of the following formula (II) was synthesized and used.
This 3-decanoyltetronic acid is an example provided with a linear alkyl group having 9 carbon atoms as R as a substituent in the general formula (2) of the above embodiment.

(Method for producing the compound of Example 2 in the synthesis method (A))
In the same production method as in the synthesis method (A) of Example 1 above, tetronic acid was synthesized as a substrate.

(Method for producing the compound of Example 2 in the synthesis method (B))
Tetronic acid (100 mg, 1.00 mmol), decanoic acid (190 mg, 1.10 mmol 1.1 equiv.), 4- (dimethylamino) pyridine (183 mg, 1.50 mmol 1.5 equiv.) Were dissolved in dichloromethane (4 mL). N- [3- (Dimethylamino)] propyl-N′-ethylcarbodiimide (290 mg, 1.50 mmol 1.5 equiv.) Was added at room temperature.
After stirring at room temperature for 12 hours, 5% aqueous citric acid solution (10 mL) was added, and the mixture was extracted with ethyl acetate (20 mL).
The resulting organic layer was separated and dried over anhydrous sodium sulfate.
The solution after drying was concentrated and purified by silica gel column chromatography using a mixed solvent of ethyl acetate-hexane to obtain 3-decanoyltetronic acid (145 mg, 57%) as a colorless oily substance.

Regarding 3-decanoyltetronic acid obtained by the synthesis method (A) or (B) of Example 2, the following results were obtained by commercially available NMR.
¹ H NMR (300 MHz, CDCl 3) δ = 0.86 (t, 3H, J = 6.9 Hz), 1.10-1.42 (m, 14H), 1.60-1.78 (m, 2H) ), 2.90 (t, 2H, J = 7.6 Hz), 4.64 (s, 1H).

Example 3
As 3-acyl-4-hydroxycoumarins, 3-decanoyl-4-hydroxycoumarin of the following formula (III) was synthesized and used.
This 3-decanoyl-4-hydroxycoumarin is an example provided with a linear alkyl group having 9 carbon atoms as R as a substituent in the general formula (3) of the above embodiment.

(Method for producing the compound of Example 3 in the synthesis method (A))
4-hydroxycoumarin was synthesized as a substrate by the same production method as the synthesis method (A) of Example 1 and Example 2 above.

(Method for producing the compound of Example 3 in the synthesis method (B))
4-hydroxycoumarin (100 mg, 0.62 mmol), decanoic acid (117 mg, 0.68 mmol 1.1 equiv.), 4- (dimethylamino) pyridine (114 mg, 0.93 mmol 1.5 equiv.) In dichloromethane ( 4 mL) and N- [3- (dimethylamino)] propyl-N′-ethylcarbodiimide (180 mg, 0.93 mmol 1.5 equiv.) Was added at room temperature.
After stirring at room temperature for 12 hours, 5% aqueous citric acid solution (10 mL) was added, and the mixture was extracted with ethyl acetate (20 mL).
The resulting organic layer was separated and dried over anhydrous sodium sulfate.
The solution after drying was concentrated and purified by silica gel column chromatography using a mixed solvent of ethyl acetate-hexane to obtain 3-decanoyl-4-hydroxycoumarin (170 mg, 87%) as a white solid.

Regarding 3-decanoyltetronic acid obtained by the synthesis method (A) or (B) of Example 3, the following results were obtained by commercially available NMR.
¹ H NMR (300 MHz, CDCl 3) δ = 0.87 (t, 3H, J = 6.9 Hz), 1.10-1.44 (m, 14H), 1.62-1.78 (m, 2H) 3.18 (t, 2H, J = 7.5 Hz), 7.25-7.38 (m, 2H), 7.67 (dt, 1H, J = 1.8, 8.1 Hz), 8. 04 (dd, 1H, J = 1.8, 8.1 Hz).

Example 4
As 3-acyl-4-hydroxycoumarins, 3-myristylinoyl-4-hydroxycoumarin of the following formula (IV) was synthesized and used.
This 3-myristylinoyl-4-hydroxycoumarin is an example provided with a linear alkyl group having 13 carbon atoms as R as a substituent in the general formula (3) of the above-described embodiment.

(Production Method of Example 4 Compound in Synthesis Method (A))
4-hydroxycoumarin was synthesized as a substrate by the same production method as the synthesis method (A) of Example 1, Example 2, and Example 3 above.

(Method for producing the compound of Example 4 in the synthesis method (B))
4-hydroxycoumarin (100 mg, 0.62 mmol), myristic acid (155 mg, 0.68 mmol 1.1 equiv.), 4- (dimethylamino) pyridine (114 mg, 0.93 mmol 1.5 equiv.) In dichloromethane (4 mL) ) And N- [3- (dimethylamino)] propyl-N′-ethylcarbodiimide (180 mg, 0.93 mmol 1.5 equiv.) Was added at room temperature.
After stirring at room temperature for 12 hours, 5% aqueous citric acid solution (10 mL) was added, and the mixture was extracted with ethyl acetate (20 mL).
The resulting organic layer was separated and dried over anhydrous sodium sulfate.
The solution after drying was concentrated and purified by silica gel column chromatography using a mixed solvent of ethyl acetate-hexane to obtain 3-myristylinoyl-4-hydroxycoumarin (173 mg, 75%) as a white solid.

Regarding 3-myristylinoyl-4-hydroxycoumarin by the synthesis method (A) or (B) of Example 4, the following results were obtained by commercially available NMR.
¹ H NMR (300 MHz, CDCl 3) δ = 0.86 (t, 3H, J = 6.9 Hz), 1.08-1.47 (m, 22H), 1.58-1.72 (m, 2H) 3.14 (t, 2H, J = 7.5 Hz), 7.25-7.38 (m, 2H), 7.66 (dt, 1H, J = 1.8, 8.1 Hz), 8. 02 (dd, 1H, J = 1.8, 8.1 Hz).

[Antimicrobial activity evaluation]
(In vitro micro liquid dilution method)
The compounds of Examples 1, 2, 3, and 4 were evaluated for antibacterial activity by an in vitro micro liquid dilution method. In addition, vancomycin was used as Comparative Example 1.
This in vitro micro liquid dilution method was performed according to the micro liquid dilution method indicated by the standard method of the Japanese Society of Chemotherapy (Japanese Society of Chemotherapy “MIC measurement method by micro liquid dilution method (micro liquid dilution method)”, Chemotherapy, 1990, 38, p. 102-105, and the Japanese Society of Chemotherapy, “Partial correction of MIC measurement method by micro liquid dilution method (Japanese Chemotherapy Standard method)”, Chemotherapy, 1993, 41, p. 189).
The results are shown in Table 1 below:

Here, the smaller the MIC value (mg / mL) in Table 1, the higher the antibacterial activity. That is, the minimum antibacterial substance concentration that inhibits bacterial growth in vitro is referred to as a MIC (minimum inhibitory concentration) value.
As a result, the compounds of Examples 1 and 2 are less active against S. aureus and E. coli, but C.I. It showed high activity against Clostridium genus including difficile and Clostridium perfringens, and bacterial selectivity was observed. In particular, the compound of Example 1 is C.I. It had activity over vancomycin against difficile. In addition, the compound of Example 2 has little effect on other bacteria, and C.I. It showed significantly higher activity than against difficile. As a result, the effect on other bacteria in the intestinal flora is reduced. It is possible to provide an antibacterial agent that can be used for treatment by infection with difficile or Clostridium perfringens.
The compounds of Examples 3 and 4 also had high activity against S. aureus. It was found to have higher activity against difficile.

(In vitro agar dilution method)
Next, antibacterial activity evaluation by in vitro agar dilution method was performed on a wide range of intestinal bacterial strains using the compounds of Examples 1 to 3 described above. The antibacterial activity evaluation by this in vitro agar dilution method was performed based on the measurement method shown by the standard method of the Japanese Society of Chemotherapy (Japanese Society of Chemotherapy “MIC method by the agar plate dilution method”, Japanese Society of Chemotherapy, Vol. 56, 2008 No. 1 (January), p. 49-57).
For the agar dilution method, ABCM agar medium (manufactured by Eiken) was used at 37 ° C. for 20 hours, and anaerobic bacteria were anaerobically cultured.
The results are shown in Table 2 below:

The MIC values in Table 2 are the same as those in the in vitro micro liquid dilution method described above. However, it is known that the in vitro agar dilution method has a relatively higher number than the in vitro micro liquid dilution method.
As a result, the same tendency as the in vitro micro liquid dilution method described above was obtained. That is, the compounds of Examples 1 and 2 are C.I. Bacterial selectivity against Clostridium bacteria including difficile and Clostridium perfringens was evident. In particular, the compound of Example 2 is C.I. There was selectivity for difficile.
In addition, the compound of Example 3 showed moderate activity overall against enterobacterial strains. It had high activity against difficile.

  Note that the configuration and operation of the above-described embodiment are examples, and it is needless to say that the configuration and operation can be appropriately changed and executed without departing from the gist of the present invention.

  The antibacterial agent of the present invention can be used for infectious diseases with few treatment methods by using a compound selective for Clostridium bacteria, and can be used industrially.

Claims (10)

A clostridium genus comprising a compound represented by 3-acyl-4-hydroxy-6-methylpyrones, a compound represented by 3-acyltetronic acids, and / or a compound represented by 3-acyl-4-hydroxycoumarins (Clostridium) Antibacterial agent for bacteria. The antibacterial agent for genus Clostridium according to claim 1, wherein the genus Clostridium is Clostridium difficile and / or Clostridium perfringens. The 3-acyl-4-hydroxy-6-methylpyrones have the formula

(Wherein R is a linear or branched chain having 4 to 30 carbon atoms or an alkyl group which may form a ring)
The antibacterial agent for Clostridium bacteria of Claim 1 or 2 characterized by the above-mentioned. The 3-acyltetronic acids have the formula

(Wherein R is a linear or branched chain having 4 to 30 carbon atoms or an alkyl group which may form a ring)
The antibacterial agent for clostridium bacteria according to any one of claims 1 to 3. The 3-acyl-4-hydroxycoumarins have the formula

(Wherein R is a linear or branched chain having 4 to 30 carbon atoms or an alkyl group which may form a ring)
The antibacterial agent for Clostridium bacteria of any one of Claims 1 thru | or 4 characterized by the above-mentioned. A composition for treating colitis for Clostridium bacteria, characterized in that the antibacterial agent according to any one of claims 1 to 5 is used. Administering a compound represented by 3-acyl-4-hydroxy-6-methylpyrones, a compound represented by 3-acyltetronic acids, and / or a compound represented by 3-acyl-4-hydroxycoumarins. A method for treating colitis in a non-human animal characterized by the above.   A compound represented by 3-acyl-4-hydroxy-6-methylpyrones, a compound represented by 3-acyltetronic acids, and / or a compound represented by 3-acyl-4-hydroxycoumarins is selected from the genus Clostridium ( Clostridium) A method for use as an antibacterial agent for bacteria. One kind of 4-hydroxy-6-methylpyrone, tetronic acid, and / or 4-hydroxycoumarin is used as a substrate, and acid chloride or acid anhydride is reacted with the substrate in the presence of an amine base in an organic solvent. The mixture is stirred at -50 ° C for 12-24 hours to synthesize a compound represented by 3-acyl-4-hydroxy-6-methylpyrones, 3-acyltetronic acids, and / or 3-acyl-4-hydroxycoumarins. The manufacturing method of the antibacterial agent for Clostridium bacteria characterized by the above-mentioned. Using 4-hydroxy-6-methylpyrone, tetronic acid, and / or 4-hydroxycoumarin as a substrate, reacting the carboxylic acid with the substrate in an organic solvent in the presence of a condensing agent and an amine base, the reaction is carried out at 0 to 50 ° C. Clostridial, characterized by synthesizing a compound represented by 3-acyl-4-hydroxy-6-methylpyrones, 3-acyltetronic acids, and 3-acyl-4-hydroxycoumarins by stirring for 12 to 24 hours A method for producing an antibacterial agent for Clostridium bacteria.