JP2005225783A - New epidithiodiketopiperadine-based antibacterial antibiotic mmrc03-0001 produced by sepedonium chlorinum - Google Patents
New epidithiodiketopiperadine-based antibacterial antibiotic mmrc03-0001 produced by sepedonium chlorinum Download PDFInfo
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本発明は、セペドニウム属に属する微生物が生産する新規エピジチオジケトピペラジン系抗生物質又はその薬理学上許容される塩、該化合物の製造方法、及び該化合物を有効成分とする抗菌剤等に関する。 The present invention relates to a novel epidithiodiketopiperazine antibiotic produced by a microorganism belonging to the genus Cepedonium or a pharmacologically acceptable salt thereof, a method for producing the compound, an antibacterial agent comprising the compound as an active ingredient, and the like.
従来、微生物が生産する抗生物質には、ペニシリン、セファロスポリン、カルバペネム等のβラクタム系抗生物質、エリスロマイシン、ジョサマイシン、ロキタマイシン等のマクロライド系抗生物質、カナマイシン、ゲンタマイシン、トブラマイシン等のアミノグコシド系抗生物質など種々の抗生物質が知られている。これらの抗生物質及びそれらを化学合成的手法により改良した化合物は、臨床上非常に有用な抗生物質として利用されてきた。しかし、これらの抗生物質は種々の感染症の治療に高頻度で用いられているため、それらの抗生物質に耐性を示す病原菌が出現して、問題化している。そこで、この病原菌の耐性の問題を解決するために、及び、更により有効で安全な抗生物質を提供するために、新しい抗菌活性等を有する抗生物質の開発が求められている。 Conventionally, antibiotics produced by microorganisms include β-lactam antibiotics such as penicillin, cephalosporin and carbapenem, macrolide antibiotics such as erythromycin, josamycin and rokitamicin, and aminogcoside antibiotics such as kanamycin, gentamicin and tobramycin Various antibiotics are known. These antibiotics and compounds obtained by improving them by chemical synthesis techniques have been utilized as clinically very useful antibiotics. However, since these antibiotics are frequently used in the treatment of various infectious diseases, pathogenic bacteria that are resistant to these antibiotics have emerged and become problematic. Therefore, in order to solve the problem of resistance of the pathogenic bacteria and to provide an even more effective and safe antibiotic, development of an antibiotic having new antibacterial activity and the like is required.
各種、抗生物質が知られている中で、エピジチオ構造を有するエピジチオジケトピペラジン系抗生物質が知られている。例えば、グリオクラディウム・ビレンス（Gliocladium・virens）等の微生物が産生する抗菌活性を有するグリオトキシン等（Journal of General Microbiology,134:2067-2075,1988；特開平７−２２７２９３号公報）や、ディコトモマイセス・シェジピィ(Dichotomomyces cejpii)等の微生物が産生する、グリオトキシンのアセチル誘導体に相当するアセチルグリオトキシン（J. Amer. Chem. Soc.,75:2110-2112,1953）等が知られている。そして、エピジチオジケトピペラジン系抗生物質の中で、アセチルグリオトキシンについては、その抗腫瘍活性も見い出されており（ＷＯ９８／２４９２６）、該化合物のアセチル基のアルコオキシ基、アシルオキシ基等の置換誘導体が合成されている。
Journal of General Microbiology,134:2067-2075,1988。 Journal of General Microbiology, 134: 2067-2075,1988.
J. Amer. Chem. Soc.,75:2110-2112,1953。 J. Amer. Chem. Soc., 75: 2110-2112, 1953.
本発明の課題は、微生物が生産し、医薬用、動物薬用、水産用、又は農園芸用薬剤として使用可能な、抗菌活性を有する新規抗生物質又はその薬理学上許容される塩、更には、該化合物の製造方法、及び該化合物を有効成分とする抗菌剤等を提供することにある。 An object of the present invention is to produce a novel antibiotic having antibacterial activity or a pharmacologically acceptable salt thereof, which is produced by a microorganism and can be used as a pharmaceutical, veterinary, marine, or agricultural or horticultural agent. It is in providing the manufacturing method of this compound, and the antibacterial agent etc. which use this compound as an active ingredient.
本発明者は、上記課題を解決すべく、鋭意探索の結果、本発明者が新しく天然界から分離した、セペドニウム属（Sepedonium属）に属する菌株が、グラム陽性細菌、特にミクロコッカス・ルテウス(Micrococcus luteus) ＡＴＣＣ９３４３のようなグラム陽性細菌に対する抗細菌活性を有する新規抗生物質を生産することを見い出し、本発明を完成するに至った。 In order to solve the above problems, the present inventor, as a result of earnest search, a strain belonging to the genus Sepedonium newly isolated from the natural world by the present inventor is a gram-positive bacterium, in particular, Micrococcus luteus (Micrococcus luteus) has been found to produce novel antibiotics with antibacterial activity against Gram-positive bacteria such as ATCC 9343 and has led to the completion of the present invention.
すなわち、本発明者は天然界の植物標本から数多くの微生物を単離し、それらの生産物について種々研究を行った結果、神奈川県秦野市弘法山にて採集したイグチの仲間の子実体より分離した糸状菌は、抗菌性を有する新規な抗細菌抗生物質を生産することを見い出した。該新規な抗細菌抗生物質を、ＭＭＲＣ０３−０００１と命名した。ＭＭＲＣ０３−０００１抗生物質は、エピジチオ構造を有するエピジチオジケトピペラジン系の新規抗生物質で、該抗生物質の化学構造は、以下の式（１）で表される。 That is, the present inventor isolated many microorganisms from plant specimens in the natural world and conducted various studies on their products, and as a result, separated from the fruit bodies of Iguchi's fellows collected at Kohoyama, Kanno City, Kanagawa Prefecture. Filamentous fungi have been found to produce new antibacterial antibiotics with antibacterial properties. The novel antibacterial antibiotic was named MMRC03-0001. The MMRC03-0001 antibiotic is an epidithiodiketopiperazine novel antibiotic having an epidithio structure, and the chemical structure of the antibiotic is represented by the following formula (1).
本発明の抗生物質ＭＭＲＣ０３−０００１を生産する微生物は、セペドニウム属に属する新規な菌株で、セペドニウム・クロリヌム（Sepedonium chlorinum）TAMA７３株と命名され、該菌株は、独立行政法人産業技術総合研究所 特許生物寄託センターにＦＥＲＭ Ｐ−１９６１７として寄託された。本発明の抗生物質ＭＭＲＣ０３−０００１を製造するには、本発明の抗生物質ＭＭＲＣ０３−０００１を生産する能力を有する微生物を培養し、培養物から該抗生物質ＭＭＲＣ０３−０００１を採取することによって製造することができる。本発明の抗生物質は、特に、ミクロコッカスのようなグラム陽性細菌に対して強い抗細菌活性を有し、医薬用、動物薬用、水産用、或いは農園芸用等の広い範囲の抗菌剤の有効成分として用いることができる。 The microorganism that produces the antibiotic MMRC03-0001 of the present invention is a new strain belonging to the genus Sepedonium, and is named Sepedonium chlorinum TAMA73 strain, which is a patent organism of the National Institute of Advanced Industrial Science and Technology. Deposited at the deposit center as FERM P-19617. In order to produce the antibiotic MMRC03-0001 of the present invention, a microorganism having the ability to produce the antibiotic MMRC03-0001 of the present invention is cultured, and the antibiotic MMRC03-0001 is collected from the culture. Can do. The antibiotic of the present invention has a strong antibacterial activity especially against gram-positive bacteria such as micrococcus, and is effective for a wide range of antibacterial agents such as pharmaceuticals, veterinary drugs, marine products, agricultural and horticultural use. It can be used as a component.
すなわち具体的には本発明は、確定したクレームを記載します。からなる。 That is, the present invention specifically describes a confirmed claim. Consists of.
また本発明は、確定したクレームを記載します。からなる。 In addition, the present invention describes a confirmed claim. Consists of.
本発明の新規抗生物質は、特に、ミクロコッカスのようなグラム陽性細菌に対して強い抗細菌活性を有し、医薬用、動物薬用、水産用、或いは農園芸用等の広い範囲に有効な抗菌剤として用いることができる。また、本発明の新規抗生物質は、本発明の菌株を用い、簡便かつ効率の良い方法で製造することが可能である。 The novel antibiotics of the present invention have a strong antibacterial activity especially against gram-positive bacteria such as micrococcus, and are effective for a wide range of medicines, veterinary drugs, fisheries, agricultural and horticultural use. It can be used as an agent. Further, the novel antibiotic of the present invention can be produced by a simple and efficient method using the strain of the present invention.
本発明は、グラム陽性細菌に対して強い抗細菌活性を有するエピジチオジケトピペラジン系の新規抗生物質ＭＭＲＣ０３−０００１、及び、その薬理学上許容される塩からなる。該本発明の抗生物質ＭＭＲＣ０３−０００１を取得するには、セペドニウム属に属する該本発明の抗生物質ＭＭＲＣ０３−０００１を生産する能力を有する微生物を培養し、培養物から該抗生物質ＭＭＲＣ０３−０００１を採取することによって行うことができる。 The present invention comprises a novel epidithiodiketopiperazine antibiotic MMRC03-0001 having strong antibacterial activity against gram-positive bacteria, and pharmacologically acceptable salts thereof. In order to obtain the antibiotic MMRC03-0001 of the present invention, a microorganism having the ability to produce the antibiotic MMRC03-0001 of the present invention belonging to the genus Sepedonium is cultured, and the antibiotic MMRC03-0001 is collected from the culture Can be done.
（本発明で使用する微生物）
本発明で、本発明の抗生物質を製造するために使用する微生物は、天然界の植物標本から数多くの微生物を単離し、それらの生産物について種々研究を行った結果、抗菌性を有する本発明の新規な抗細菌抗生物質ＭＭＲＣ０３−０００１を生産することを見い出したことにより取得した新規な菌株であり、以下のように同定された。
(Microorganism used in the present invention)
In the present invention, the microorganisms used for producing the antibiotics of the present invention are isolated from a large number of microorganisms from natural plant specimens, and as a result of various studies on their products, the present invention has antibacterial properties. This is a novel strain obtained by finding that it produces MMRC03-0001, a novel antibacterial antibiotic, and was identified as follows.
本発明の抗生物質ＭＭＲＣ０３−０００１を生産するＴＡＭＡ７３株は、次のような菌学的性状を有する：麦芽エキス寒天、三浦培地、オートミール寒天などの上において生育は早く拡散し、２５℃、７日間でコロニーは６０ｍｍに達するが、その菌叢は疎らで、綿毛状、厚壁胞子を形成する場合は表面、裏面とも鮮黄色を呈する。分生子柄は気菌糸上に形成され、フィアライドは規則正しく数個の輪生状に形成、１１．９−７０．６×２．３−４．０μｍ（平均４３．４×３．０μｍ）、Ｌ／Ｗ ３．８−２４．７（平均１４．５）、先端は直径１．９−２．３μｍに細くなり不明瞭なカラーを有し分生子を塊状に形成し、分生子は楕円形、円筒形、倒卵形など様々で、無色、平滑、１．９−１３．４×１．４−４．５μｍ（平均５．６×２．１μｍ）、Ｌ／Ｗ １．０４−６．１８（平均２．７７）、厚壁胞子は気菌糸あるいは基生菌糸に側生する無色の矩形細胞上に生じ、倒卵形、ボート形あるいは先端がとがった楕円形、黄金色で、顕著な縦の縞模様をもち、３４．７−５１．７×１４．５−２１．５μｍ（平均４２．８×１７．４μｍ）、Ｌ／Ｗ ２．１０−３．０４（平均２．４８）となる。 The TAMA73 strain producing the antibiotic MMRC03-0001 of the present invention has the following mycological characteristics: growth spreads quickly on malt extract agar, Miura medium, oatmeal agar, etc., 25 ° C., 7 days The colony reaches 60 mm, but the flora is sparse, and when forming fluffy and thick-wall spores, the front and back surfaces are bright yellow. Conidia are formed on aerial hyphae, and phialides are regularly formed into several ring-like shapes, 11.9-70.6 × 2.3-4.0 μm (average 43.4 × 3.0 μm), L / W 3.8-24.7 (average 14.5), the tip is 1.9-2.3 μm in diameter, has an unclear color and forms conidia in a lump shape, the conidia is oval, Various shapes such as cylindrical shape and egg shape, colorless, smooth, 1.9-13.4 × 1.4-4.5 μm (average 5.6 × 2.1 μm), L / W 1.04-6.18 (average) 2.77), thick-wall spores are formed on colorless rectangular cells lateral to the aerial hyphae or basal hyphae, and fallen oval, boat-shaped or elliptical with a pointed tip, golden color, and a remarkable vertical stripe pattern. 34.7-51.7 × 14.5-21.5 μm (average 42.8 × 17.4 μm), L / W 2.10-3.04 (flat 2.48) and a.
本菌株は、担子菌子実体上より得られ、生育が早く、綿毛状、鮮黄色で、無色の規則正く輪生するVerticillium型のフィアロ型分生子と鮮黄色大型のアレウロ型分生子（厚壁胞子）を形成することからセペドニウム（Sepedonium）属に属することは明らかである。菌類寄生菌として知られる本属の中からは、倒卵形で顕著な縞模様を有する厚壁胞子を作ることから本菌は容易にセペドニウム・クロリヌム（Sepedonium chlorinum ）(Tul. & C. Tul.) Damonに属することがわかる。そこで本菌株をセペドニウム・クロリヌム(Tul. & C. Tul.) Damon ＴＡＭＡ７３と同定した。なお本菌株は独立行政法人産業技術総合研究所特許生物寄託センターにＦＥＲＭ Ｐ−１９６１７として寄託されている。 This strain is obtained from a basidiomycete fruit body, grows fast, fluffy, bright yellow, colorless and regularly rotatively verticillium type fiaro type conidia and bright yellow large allelo type conidia (thick wall) It is clear that it belongs to the genus Sepedonium from the formation of spores. Among the genera known as fungal parasites, they produce a thick-walled spore with an inverted egg shape and striking stripes, which makes it easy for Sepedonium chlorinum (Tul. & C. Tul.) Damon It is understood that it belongs to. Therefore, this strain was identified as Cepedonium chlorinum (Tul. & C. Tul.) Damon TAMA73. This strain has been deposited as FERM P-19617 at the National Institute of Advanced Industrial Science and Technology Patent Organism Depositary.
ＴＡＭＡ７３株は、一般の糸状菌同様、変異しやすい性質を有している。例えば、ＴＡＭＡ７３株あるいはこの株由来の突然変異株、形質接合株、遺伝子組換体であってもＭＭＲＣ０３−０００１を生産する菌株はすべて本発明に使用できる。またセペドニウム・クロリヌムのテレオモルフはヒポミセス・クロリゲヌム（Hypomyces chlorinigenus） Rogerson & Samuelsであるとされている。そこで、セペドニウムのみならずヒポミセス属菌株でも本物質を生産する菌株はすべて本発明に使用できる。 The TAMA73 strain has the property of being easily mutated, like general filamentous fungi. For example, all strains producing MMRC03-0001 can be used in the present invention, even the TAMA73 strain or a mutant strain, a zygote strain, or a gene recombinant derived from this strain. In addition, the teleomorph of Sepedonium chlorinum is said to be Hypomyces chlorinigenus Rogerson & Samuels. Therefore, not only cepedonium but also strains of the genus Hipomyces can be used in the present invention.
（本発明の微生物の培養、抗生物質の採取）
本発明で、抗生物質ＭＭＲＣ０３−０００１を製造するには、セペドニウム属に属し、抗生物質ＭＭＲＣ０３−０００１を生産する能力を有する本発明の微生物を培養し、培養物中に抗生物質ＭＭＲＣ０３−０００１を生産、蓄積する。ＭＭＲＣ０３−０００１生産菌の培養には、通常の微生物が利用できる栄養物を含有する培地が利用できる。栄養源としては、従来糸状菌の培養に利用されている公知のものが使用できる。たとえば、炭素源としてはブドウ糖、蔗糖、グリセリン、澱粉、麦芽糖、乳糖、糖蜜、動植物油脂、デキストリンなどが利用可能である。窒素源としては大豆粉、小麦胚芽、コーンスティープリカー，綿実粕，肉エキス、酵母エキス、ペプトン、硝酸態あるいはアンモニウム態窒素などが利用可能である。必要に応じ、塩類、金属イオンを含む無機塩類を添加することが可能である。また生産菌の生育あるいはＭＭＲＣ０３−０００１の生産を向上させるために有機あるいは無機物を添加することができる。
(Culture of microorganisms of the present invention, collection of antibiotics)
In order to produce the antibiotic MMRC03-0001 in the present invention, the microorganism of the present invention belonging to the genus Sepedonium and capable of producing the antibiotic MMRC03-0001 is cultured, and the antibiotic MMRC03-0001 is produced in the culture. ,accumulate. For the culture of MMRC03-0001 producing bacteria, a medium containing nutrients that can be used by ordinary microorganisms can be used. As the nutrient source, known ones conventionally used for culturing filamentous fungi can be used. For example, glucose, sucrose, glycerin, starch, maltose, lactose, molasses, animal and vegetable oils and fats, dextrin and the like can be used as the carbon source. As the nitrogen source, soybean flour, wheat germ, corn steep liquor, cottonseed meal, meat extract, yeast extract, peptone, nitrate or ammonium nitrogen can be used. If necessary, it is possible to add salts and inorganic salts containing metal ions. Organic or inorganic substances can be added to improve the growth of the producing bacteria or the production of MMRC03-0001.
培養法としては好気的条件での培養法、固体基質上での固体静置培養あるいは液体培地による深部培養が可能である。培養に適当な温度は１０〜３２℃であるが、通常は２０〜３０℃で培養する。培養日数は培養方法と培地の種類によっても異なるが、数日から数１０日で目的物質ＭＭＲＣ０３−０００１は生産される。
抗生物質ＭＭＲＣ０３−０００１の検定にあたっては以下の方法が用いられる。すなわち、培養液からｎ−ブタノールなどの有機溶媒粗抽出液中のＭＭＲＣ０３−０００１或いは精製されたＭＭＲＣ０３−０００１は病原細菌ミクロコッカス・ルテウスＡＴＣＣ ９３４３を被検菌とした生物学的方法により検出される。或いは、Ｃ１８逆層カラムを接続し、フォトダイオードアレー検出器を装着した高速液体クロマトグラフィーさらには質量分析計を直結した高速液体クロマトグラフ・質量分析装置などの化学的方法によってもＭＭＲＣ０３−０００１の検出を行うことができる。
As a culture method, a culture method under an aerobic condition, a solid stationary culture on a solid substrate, or a deep culture using a liquid medium is possible. A suitable temperature for culturing is 10 to 32 ° C., but usually culturing at 20 to 30 ° C. The number of culture days varies depending on the culture method and the type of medium, but the target substance MMRC03-0001 is produced in several days to several tens of days.
The following method is used for testing antibiotic MMRC03-0001. That is, MMRC03-0001 or purified MMRC03-0001 in an organic solvent crude extract such as n-butanol is detected from a culture solution by a biological method using the pathogenic bacterium Micrococcus luteus ATCC 9343 as a test bacterium. . Alternatively, MMRC03-0001 can also be detected by chemical methods such as high-performance liquid chromatography connected to a C18 reverse layer column and equipped with a photodiode array detector, or a high-performance liquid chromatograph / mass spectrometer directly connected to a mass spectrometer. It can be performed.
抗生物質ＭＭＲＣ０３−０００１を培養液から採取するにあたっては活性炭、アルミナ、シリカゲルなどの吸着剤、ゲルろ過剤などが使用されうるが、次に示す方法がより効果的である：培養液から目的物をｎ−ブタノールで抽出し、ｎ−ブタノール層を減圧下で濃縮することにより、ＭＭＲＣ０３−０００１を含有するオイル状物質を得る。更に、目的物以外の物質を除くため、以下に示す前処理を行うことができる。すなわち、ｎ−ブタノール濃縮物に水を加えた後、酢酸エチルで再抽出し、得られた酢酸エチル層を再び減圧下で濃縮してオイル状物質を得る。 In collecting the antibiotic MMRC03-0001 from the culture solution, an adsorbent such as activated carbon, alumina and silica gel, a gel filtration agent and the like can be used, but the following method is more effective: By extracting with n-butanol and concentrating the n-butanol layer under reduced pressure, an oily substance containing MMRC03-0001 is obtained. Furthermore, in order to remove substances other than the target product, the following pretreatment can be performed. That is, water is added to the n-butanol concentrate, followed by re-extraction with ethyl acetate, and the resulting ethyl acetate layer is again concentrated under reduced pressure to obtain an oily substance.
更に、酢酸エチル濃縮物に９０％メタノール溶液を加え、等倍容のｎ−ヘキサンを加え、撹拌後、９０％メタノール溶液を集める。更に、９０％メタノール溶液層を７５％メタノール溶液となるよう水を加えた後、等倍容の四塩化炭素を加え、撹拌後、７５％メタノール溶液層を集め、減圧下で濃縮することにより目的物を含む固形物を得る。これをシリカゲルのカラムに吸着させ、ｎ−ヘキサン、アセトン、酢酸エチル、メタノールなどの有機溶媒の混合液で目的物の溶出を行う。目的物はさらに種々のクロマト担体と溶媒を組み合わせて薄層クロマトグラフ、低圧カラムクロマトグラフ、高速液体クロマトグラフなどを行い、ＭＭＲＣ０３−０００１を単離する。単離された本化合物の構造は化合物の種々なスペクトル特性、特にその核磁気共鳴、質量、紫外部、赤外部スペクトルを詳細な解析により決定される。 Furthermore, a 90% methanol solution is added to the ethyl acetate concentrate, and an equal volume of n-hexane is added. After stirring, the 90% methanol solution is collected. Further, after adding water so that the 90% methanol solution layer becomes a 75% methanol solution, an equal volume of carbon tetrachloride is added, and after stirring, the 75% methanol solution layer is collected and concentrated under reduced pressure. A solid containing the product is obtained. This is adsorbed on a silica gel column, and the target product is eluted with a mixed solution of organic solvents such as n-hexane, acetone, ethyl acetate, and methanol. The target product is further subjected to thin layer chromatography, low pressure column chromatography, high performance liquid chromatography, etc. by combining various chromatographic carriers and solvents to isolate MMRC03-0001. The structure of the isolated compound is determined by detailed analysis of the various spectral properties of the compound, particularly its nuclear magnetic resonance, mass, ultraviolet and infrared spectra.
（ＭＭＲＣ０３−０００１の理化学的性質）
抗生物質ＭＭＲＣ０３−０００１は次の理化学的性質を有する。
１）分子量：３２６
２）マススペクトル：FAB-MS(NBA)；m/z 349[M+Na]+, 327[M+H]+, 262[M-2S]+
分子式（C13H14N2O4S2）
３）紫外線吸収スペクトル：(max nm(() in MeOH; 232 weak shoulder (4270)
４）赤外線吸収スペクトル：(max cm-1 in KBr; 3360, 1680, 1670, 1400, 1 360, 1060, 1020, 875, 740, 620
５）核磁気共鳴スペクトル：1H-NMR(400 MHz, CDCl3) ( (ppm): 2.00 (3H, s ), 2.83 (1H, m), 3.08 (3H, s), 3.60 (1H, m), 3.6 5 (1H, m), 4.38 (1H, m), 4.44 (1H, m), 4.63 (1H, m), 5.96 (1H, m)
13C-NMR(100.5 MHz, CDCl3) ( (ppm): 18.3, 27.6, 37.7, 53.7, 55. 9, 56.9, 62.2, 73.6, 74.2, 124.2, 134.4, 163.0, 165.8
６）旋光度：[a] D −２４０°(c=0.38, CHCl3)
７）溶解性：クロロホルム、酢酸エチル、アセトニトリル、アセトン、メタノー ル、エタノール、ジメチルスルホキシドに易溶、n-ヘキサン、四塩 化炭素、水に難溶
８）化学構造式：前述の通り
（ＭＭＲＣ０３−０００１の抗菌剤としての利用）
本発明の抗生物質ＭＭＲＣ０３−０００１は、医薬用、動物薬用、水産用、或いは農園芸用等の広い範囲の抗菌剤として用いることができる。
本発明の抗生物質を抗菌剤の有効成分として用いる場合は、本発明のＭＭＲＣ０３−０００１生産菌株を培養して得た培養物から、本発明のＭＭＲＣ０３−０００１を上記のような手段で、精製して用いることができる。また、抗菌剤の利用分野によっては、粗精製物の状態で用いることもできる。
(Physicochemical properties of MMRC03-0001)
Antibiotic MMRC03-0001 has the following physicochemical properties:
1) Molecular weight: 326
2) Mass spectrum: FAB-MS (NBA); m / z 349 [M + Na] + , 327 [M + H] + , 262 [M-2S] +
Molecular formula (C 13 H 14 N 2 O 4 S 2 )
3) UV absorption spectrum: ( max nm (() in MeOH; 232 weak shoulder (4270)
4) Infrared absorption spectrum: ( max cm -1 in KBr; 3360, 1680, 1670, 1400, 1 360, 1060, 1020, 875, 740, 620
5) Nuclear magnetic resonance spectrum: 1 H-NMR (400 MHz, CDCl 3 ) ((ppm): 2.00 (3H, s), 2.83 (1H, m), 3.08 (3H, s), 3.60 (1H, m) , 3.6 5 (1H, m), 4.38 (1H, m), 4.44 (1H, m), 4.63 (1H, m), 5.96 (1H, m)
13 C-NMR (100.5 MHz, CDCl 3 ) ((ppm): 18.3, 27.6, 37.7, 53.7, 55. 9, 56.9, 62.2, 73.6, 74.2, 124.2, 134.4, 163.0, 165.8
6) Optical rotation: [a] D −240 ° (c = 0.38, CHCl 3 )
7) Solubility: Soluble in chloroform, ethyl acetate, acetonitrile, acetone, methanol, ethanol, dimethyl sulfoxide, n-hexane, carbon tetrachloride, sparingly soluble in water 8) Chemical structural formula: As described above (MMRC03- Use of 0001 as an antibacterial agent
The antibiotic MMRC03-0001 of the present invention can be used as a wide range of antibacterial agents such as pharmaceuticals, veterinary drugs, fisheries, agricultural and horticultural use.
When the antibiotic of the present invention is used as an active ingredient of an antibacterial agent, MMRC03-0001 of the present invention is purified from the culture obtained by culturing the MMRC03-0001 producing strain of the present invention by the above-described means. Can be used. Further, depending on the field of use of the antibacterial agent, it can be used in the state of a roughly purified product.
本発明の抗生物質ＭＭＲＣ０３−０００１を、医薬等として用いる場合には、該化合物を、薬理学上許容される塩の形で用いることができる。かかる薬理学上許容される塩は、薬理学上公知のものを用いることができるが、例えば、薬理学上許容される金属塩、アンモニウム塩、含有アミン付加塩、アミノ酸付加塩等を用いることができる。金属塩としては、リチウム塩、ナトリウム塩、カリウム塩等のアルカリ金属塩、マグネシウム塩、カルシウム塩等のアルカリ土類金属塩、アルミニウム塩、亜鉛塩等が挙げられ、有機アミン付加塩としてはモルホリン、ピペリジン等の付加塩、アミノ酸付加塩としては、グリシン、フェニルアラニン、アスパラギン酸、グルタミン酸、リジン等の付加塩が挙げられる。本発明の抗生物質ＭＭＲＣ０３−０００１、又は、該化合物の薬理学上許容される塩を医薬として投与するには、該有効成分を、製薬上用いられる適宜の形態に製剤化して、全身又は局所的に、経口又は非経口の形で投与することができる。製剤形態としては、例えば、錠剤、散剤、顆粒剤、シロップ剤、注射剤、液剤、乳剤、懸濁剤、軟膏等を挙げることができる。該製剤化に際しては、例えば、各種の賦形剤、潤沢剤、結合剤、崩壊剤、懸濁化剤、乳化剤等の補助剤を用いることができる。 When the antibiotic MMRC03-0001 of the present invention is used as a medicine or the like, the compound can be used in the form of a pharmacologically acceptable salt. As such pharmacologically acceptable salts, those known in pharmacology can be used. For example, pharmaceutically acceptable metal salts, ammonium salts, containing amine addition salts, amino acid addition salts, and the like can be used. it can. Examples of the metal salt include alkali metal salts such as lithium salt, sodium salt and potassium salt, alkaline earth metal salts such as magnesium salt and calcium salt, aluminum salt, zinc salt and the like, and organic amine addition salt includes morpholine, Examples of addition salts such as piperidine and amino acid addition salts include addition salts such as glycine, phenylalanine, aspartic acid, glutamic acid, and lysine. In order to administer the antibiotic MMRC03-0001 of the present invention or a pharmacologically acceptable salt of the compound as a pharmaceutical, the active ingredient is formulated into an appropriate form for pharmacological use, and systemically or locally. In addition, it can be administered orally or parenterally. Examples of the dosage form include tablets, powders, granules, syrups, injections, solutions, emulsions, suspensions, ointments and the like. In the formulation, for example, various excipients, lubricants, binders, disintegrants, suspending agents, auxiliary agents such as emulsifiers can be used.
本発明の抗生物質ＭＭＲＣ０３−０００１を、農園芸用等の抗菌剤として用いる場合には、該化合物又はその塩を、適宜の補助剤とともに製剤化して、例えば、粉剤、粒剤、乳化剤、水和剤のような形で用いることができる。また、本発明の抗生物質ＭＭＲＣ０３−０００１を、動物薬用の抗菌剤として用いる場合には、該化合物又はその塩を適宜の投与形態で、動物医薬として、或いは、該薬剤を飼料等に混合した形で投与することができる。また、本発明の抗生物質ＭＭＲＣ０３−０００１を、水産用等の抗菌剤として用いる場合には、該化合物又はその塩を、慣用の希釈剤、担体、増量剤、添加剤等を用いて製剤化し、養魚用の水等の抗菌処理に用いることができる。また、該抗菌剤を養魚用飼料等に添加して用いることもできる。更に、該抗菌剤を塗布剤の形に製剤化し、魚網等の漁労用具や漁労施設の抗菌処理に用いることができる。 When the antibiotic MMRC03-0001 of the present invention is used as an antibacterial agent for agricultural or horticultural use, the compound or a salt thereof is formulated with an appropriate auxiliary agent, for example, powder, granule, emulsifier, hydration It can be used in the form of an agent. Further, when the antibiotic MMRC03-0001 of the present invention is used as an antibacterial agent for veterinary drugs, the compound or a salt thereof in an appropriate dosage form, veterinary medicine, or a form in which the drug is mixed with feed or the like Can be administered. In addition, when the antibiotic MMRC03-0001 of the present invention is used as an antibacterial agent for fisheries or the like, the compound or a salt thereof is formulated using a conventional diluent, carrier, extender, additive, etc. It can be used for antibacterial treatment such as water for fish farming. In addition, the antibacterial agent can be used by adding to fish feed and the like. Further, the antibacterial agent can be formulated in the form of a coating agent and used for antibacterial treatment of fishing equipment such as fish nets and fishing facilities.
以下、実施例により本発明をより具体的に説明するが、本発明の技術的範囲はこれらの例示に限定されるものではない。 EXAMPLES Hereinafter, although an Example demonstrates this invention more concretely, the technical scope of this invention is not limited to these illustrations.
（抗生物質ＭＭＲＣ０３−０００１の調製及び生物試験）
セペドニウム・クロリヌム ＴＡＭＡ７３株を麦芽エキス寒天斜面培地上において２５℃、１０日間培養した。この斜面培養で得られた菌糸体を２０ｍｌの生産培地（ブドウ糖２％、グリセリン３％、ポリペプトン（日本製薬）０．５％、酵母エキス（日本製薬）０．２％、食塩０．３％、炭酸カルシウム１％）を含む２５０ｍｌ容の三角フラスコ５０本に植菌して、２５℃、２２０ｒｐｍにて回転振盪培養を行った。培養液のｐＨは培養に伴い上昇し、同時にＭＭＲＣ０３−０００１の生産性も１２日でピークに到達した。こうして、約２１ｍｇのＭＭＲＣ０３−０００１を含む培養液１０００ｍｌを得た。１２日目の培養物に７５０ｍｌのｎ−ブタノールを加え撹拌後、冷蔵庫内（５℃）で一晩、保存した。その後、ｎ−ブタノールを集め、減圧下で濃縮し、２．３ｇのオイル状物質を得た。
(Preparation and biological test of antibiotic MMRC03-0001)
Cepedonium chlorinum TAMA73 strain was cultured on a malt extract agar slant medium at 25 ° C. for 10 days. The mycelium obtained by this slope culture was used in a 20 ml production medium (glucose 2%, glycerin 3%, polypeptone (Nippon Pharmaceutical) 0.5%, yeast extract (Nippon Pharmaceutical) 0.2%, salt 0.3%, The cells were inoculated into 50 250 ml Erlenmeyer flasks containing 1% of calcium carbonate, and subjected to rotary shaking culture at 25 ° C. and 220 rpm. The pH of the culture increased with the culturing, and at the same time, the productivity of MMRC03-0001 reached its peak in 12 days. Thus, 1000 ml of a culture solution containing about 21 mg of MMRC03-0001 was obtained. 750 ml of n-butanol was added to the culture on the 12th day, stirred, and stored overnight in a refrigerator (5 ° C.). Thereafter, n-butanol was collected and concentrated under reduced pressure to obtain 2.3 g of an oily substance.
そのオイル状物質に２５０ｍｌ酢酸エチルと同量の水を加え撹拌後、酢酸エチル層と水層に分けた。さらに水層は２５０ｍｌの酢酸エチルを加え、撹拌後、再び酢酸エチル層と水層に分けた。それぞれの酢酸エチル層を併せて減圧下で濃縮後、１．９ｇのオイル状物質を得た。そのオイル状物質に２５０ｍｌの９０％メタノール溶液と同量のｎ−ヘキサンを加え、撹拌後、ｎ−ヘキサン層と９０％メタノール溶液層に分けた。得られた２５０ｍｌの９０％メタノール溶液層に５０ｍｌの水を加え、更に３００ｍｌの四塩化炭素を加え、７５％メタノール溶液層と四塩化炭素に分けた。得られた３００ｍｌの７５％メタノール溶液を減圧下で濃縮後、約３００ｍｇの固形物を得た。その固形物は少量の酢酸エチルに溶かし、あらかじめシリカゲルをｎ−ヘキサンでつめたカラム（外径２．５ｃｍ×長さ２０ｃｍ）に吸着させ、ｎ−ヘキサン：酢酸エチルの系で酢酸エチルを段階的に高めていくことにより目的物質を溶出させた。この結果、ＭＭＲＣ０３−０００１を含有するフラクション４００ｍｌを得た。 The same amount of water as 250 ml ethyl acetate was added to the oily substance and stirred, and then the mixture was separated into an ethyl acetate layer and an aqueous layer. Further, 250 ml of ethyl acetate was added to the aqueous layer, and after stirring, the mixture was again divided into an ethyl acetate layer and an aqueous layer. The ethyl acetate layers were combined and concentrated under reduced pressure to obtain 1.9 g of an oily substance. The same amount of n-hexane as 250 ml of 90% methanol solution was added to the oily substance, and after stirring, it was divided into an n-hexane layer and a 90% methanol solution layer. 50 ml of water was added to the obtained 250 ml of 90% methanol solution layer, 300 ml of carbon tetrachloride was further added, and the mixture was divided into a 75% methanol solution layer and carbon tetrachloride. The obtained 300 ml of 75% methanol solution was concentrated under reduced pressure to obtain about 300 mg of a solid. The solid matter was dissolved in a small amount of ethyl acetate, adsorbed on a column (outer diameter 2.5 cm × length 20 cm) previously packed with silica gel, and stepped into ethyl acetate in an n-hexane: ethyl acetate system. The target substance was eluted by elevating it to a high level. As a result, a 400 ml fraction containing MMRC03-0001 was obtained.
そのフラクションは減圧下で濃縮後、ＭＭＲＣ０３−０００１を含む１２６ｍｇの固形物を得た。得られた固形物はクロロホルム：メタノール＝１：１の混液２ｍｌに溶かし、その１ｍｌを逆相カラム（資生堂カプセルパックＣ１８、ＳＧ１２０、５μｍ、外径２０ｍｍ×長さ２５０ｍｍ）を接続した高速液体クロマトグラフにて分取を行った。移動層溶媒にはアセトニトリル：水の系を用い、流速７ｍｌ／分でアセトニトリルを６０分間で５％から５０％へ直線的に高めることで溶出を行った。目的のＭＭＲＣ０３−０００１は４１分で溶出され、ＭＭＲＣ０３−０００１を含有する２１ｍｌのフラクションを得た。そのフラクションを減圧下で濃縮後、２６ｍｇの固形物を得た。 The fraction was concentrated under reduced pressure to obtain 126 mg of a solid containing MMRC03-0001. The obtained solid was dissolved in 2 ml of a mixture of chloroform: methanol = 1: 1, and 1 ml of the solid was connected to a reverse phase column (Shiseido Capsule Pack C18, SG120, 5 μm, outer diameter 20 mm × length 250 mm). Sorting was performed at Elution was performed by using a system of acetonitrile: water as the moving bed solvent and linearly increasing acetonitrile from 5% to 50% in 60 minutes at a flow rate of 7 ml / min. The desired MMRC03-0001 eluted at 41 minutes, yielding 21 ml fractions containing MMRC03-0001. The fraction was concentrated under reduced pressure to obtain 26 mg of a solid.
その固形物はクロロホルム：メタノール＝１：１の混液１ｍｌに溶かし、逆相カラム（資生堂カプセルパックＣ１８、ＳＧ１２０、５μｍ、外径２０ｍｍ×長さ２５０ｍｍ）を接続した高速液体クロマトグラフにより再分取を行った。移動層溶媒にはアセトニトリル：水の系を用い、流速７ｍｌ／分でアセトニトリルを６０分間で５％から３０％へ直線的に高め、３０％で３０分間保持し、その後３０分間で３０％から５０％へ直線的に高めて溶出を行った。目的のＭＭＲＣ０３−０００１は約５８分で溶出され、ＭＭＲＣ０３−０００１を含有する２１ｍｌのフラクションを得た。そのフラクションを減圧下で濃縮後、８ｍｇの無晶形白色粉末を得た。 The solid was dissolved in 1 ml of a mixed solution of chloroform: methanol = 1: 1 and re-sorted by a high performance liquid chromatograph connected with a reverse phase column (Shiseido Capsule Pack C18, SG120, 5 μm, outer diameter 20 mm × length 250 mm). went. As the moving bed solvent, acetonitrile: water system was used, and acetonitrile was linearly increased from 5% to 30% in 60 minutes at a flow rate of 7 ml / min, held at 30% for 30 minutes, and then from 30% to 50 in 30 minutes. Elution was carried out with a linear increase to%. The desired MMRC03-0001 eluted at about 58 minutes, yielding a 21 ml fraction containing MMRC03-0001. The fraction was concentrated under reduced pressure to obtain 8 mg of amorphous white powder.
得られたＭＭＲＣ０３−０００１の種々の量を含む検定培地１ｍｌに、１０4個のミクロコッカス・ルテウス ＡＴＣＣ９３４３の細胞を接種し、３７℃で２日間培養後、それぞれの濁度を測定して、ミクロコッカス・ルテウス ＡＴＣＣ９３４３に対する最小阻止濃度（ＭＩＣ100）を求めた。その結果、ＭＭＲＣ０３−０００１のミクロコッカス・ルテウス ＡＴＣＣ９３４３に対するＭＩＣ100は２．５μｇ／ｍｌであった。このように、本発明による抗菌剤ＭＭＲＣ０３−０００１はグラム陽性細菌、特にミクロコッカス・ルテウス ＡＴＣＣ９３４３に対して優れた抗菌活性を示すことから、グラム陽性細菌感染症に対する有効な治療薬、消毒薬、動物薬、水産用薬品、農薬となる。 In assay medium 1ml containing various amounts of the resulting MMRC03-0001, inoculated with 10 4 cells of Micrococcus luteus ATCC9343, after 2 days of culture at 37 ° C., by measuring the respective turbidity, micro The minimum inhibitory concentration (MIC 100 ) for Coccus luteus ATCC 9343 was determined. As a result, MIC 100 against Micrococcus luteus ATCC9343 of MMRC03-0001 was 2.5 [mu] g / ml. Thus, since the antibacterial agent MMRC03-0001 according to the present invention exhibits excellent antibacterial activity against Gram-positive bacteria, particularly Micrococcus luteus ATCC 9343, effective therapeutic agents, disinfectants, and animals for Gram-positive bacterial infections It becomes medicine, marine chemicals, and agricultural chemicals.
（抗生物質ＭＭＲＣ０３−０００１の製造）
よく生育したセペドニウム・クロリヌム ＴＡＭＡ７３株の麦芽エキス寒天斜面培養から１白金耳の菌糸体を７０ｍｌの種培地（グリセリン２％、ブドウ糖０．５％、大豆粉１％、酵母エキス０．２％、食塩０．２５％、炭酸カルシウム０．４％）を含む５００ｍｌ容の三角フラスコ５本に植菌して２５℃、２２０ｒｐｍにて３日間、回転振盪機上で培養した。得られた種培養を３５０ｍｌを５０Ｌジャー培養の種母として用いた。ジャー培養は、本培養培地３０Ｌ（グリセリン２％、ブドウ糖０．５％、大豆粉２％、酵母エキス０．２％、食塩０．２５％、炭酸カルシウム０．４％、消泡剤（ＣＣ−４３８、日本油脂）０．０３％）を含む５０Ｌジャーファーメンター１基に植菌して、２７℃、通気量１５Ｌ／ｍｉｎ(０．５ＶＶＭ)、圧力０．５ｋｇ／ｃｍ2・Ｇ、初期攪拌２００ｒｐｍにて培養を開始し、攪拌は２００〜６００ｒｐｍの間で溶存酸素量１０％以上に調節した。培養開始直後のｐＨは７付近であったが、７２時間後に８．０に到達し、菌体量はほぼ定常状態となった。１００時間培養後、ｐＨは８．５となり、７１０ｍｇのＭＭＲＣ０３−０００１を含む培養液２７Ｌを得た。
(Production of antibiotic MMRC03-0001)
Well-grown Cepedonium chlorinum TAMA73 strain malt extract agar slope culture 1 platinum ear mycelium in 70 ml seed medium (glycerin 2%, glucose 0.5%, soy flour 1%, yeast extract 0.2%, salt Five 500 ml Erlenmeyer flasks containing 0.25% and calcium carbonate 0.4%) were inoculated and cultured at 25 ° C. and 220 rpm for 3 days on a rotary shaker. 350 ml of the obtained seed culture was used as a seed mother for 50 L jar culture. Jar culture is 30L of main culture medium (glycerin 2%, glucose 0.5%, soy flour 2%, yeast extract 0.2%, salt 0.25%, calcium carbonate 0.4%, antifoam (CC- Inoculated into one 50L jar fermenter containing 438, Japanese oil and fat) 0.03%), 27 ° C, aeration rate 15L / min (0.5VVM), pressure 0.5kg / cm 2 · G, initial stirring Incubation was started at 200 rpm, and stirring was adjusted to a dissolved oxygen content of 10% or more between 200 and 600 rpm. Although the pH immediately after the start of the culture was around 7, it reached 8.0 after 72 hours, and the amount of cells was almost in a steady state. After culturing for 100 hours, the pH became 8.5, and 27 L of a culture solution containing 710 mg of MMRC03-0001 was obtained.
得られた培養液をブフナーロートで減圧濾過し、約５ｋｇのケーキと２７Ｌのロ液に分けた。得られたケーキは１０Ｌのメタノールで抽出し、メタノール溶液と菌体に濾別した。一方、培養ロ液は２７Ｌの酢酸エチルで抽出した。得られたケーキのメタノール抽出液は減圧下で濃縮後、１０Ｌの酢酸エチルと同量の水を加え撹拌後、酢酸エチル層と水層に分けた。得られた酢酸エチル層は培養ロ液の酢酸エチル抽出液と併せて、減圧下で濃縮後、約６３ｇのオイル状物質を得た。そのオイル状物質に７．５Ｌの９０％メタノール溶液と同量のｎ−ヘキサンを加え、撹拌後、ｎ−ヘキサン層と９０％メタノール溶液層に分けた。得られた７．５Ｌの９０％メタノール溶液層に１．５Ｌの水を加え、更に９Ｌの四塩化炭素を加え７５％メタノール溶液層と四塩化炭素に分けた。 The obtained culture solution was filtered under reduced pressure using a Buchner funnel, and divided into about 5 kg of cake and 27 L of filtrate. The obtained cake was extracted with 10 L of methanol and separated into a methanol solution and cells. On the other hand, the culture broth was extracted with 27 L of ethyl acetate. The methanol extract of the obtained cake was concentrated under reduced pressure, 10 L of ethyl acetate in the same amount of water was added and stirred, and then the mixture was separated into an ethyl acetate layer and an aqueous layer. The obtained ethyl acetate layer was combined with the ethyl acetate extract of the culture broth and concentrated under reduced pressure to obtain about 63 g of an oily substance. The same amount of n-hexane as 7.5 L of 90% methanol solution was added to the oily substance, and after stirring, it was divided into an n-hexane layer and a 90% methanol solution layer. 1.5 L of water was added to the obtained 7.5 L of 90% methanol solution layer, and 9 L of carbon tetrachloride was further added to separate into 75% methanol solution layer and carbon tetrachloride.
得られた９Ｌの７５％メタノール溶液を減圧下で濃縮後、約１１ｇの固形物を得た。得られた固形物は４０ｍｌの酢酸エチルに溶かし、あらかじめシリカゲルをｎ−ヘキサンでつめたカラム（外径８ｃｍ×長さ３４ｃｍ）に吸着させ、ｎ−ヘキサン：酢酸エチルの系で酢酸エチルを段階的に高めて目的物質を溶出させた。この結果、ＭＭＲＣ０３−０００１を含有するフラクション８．５Ｌを得た。そのフラクションは減圧下で濃縮後、ＭＭＲＣ０３−０００１を含む約３．８ｇの固形物を得た。 The obtained 9 L of 75% methanol solution was concentrated under reduced pressure to obtain about 11 g of a solid. The obtained solid was dissolved in 40 ml of ethyl acetate, adsorbed on a column (outer diameter 8 cm × length 34 cm) previously packed with silica gel, and stepped with ethyl acetate in a system of n-hexane: ethyl acetate. And the target substance was eluted. As a result, a 8.5 L fraction containing MMRC03-0001 was obtained. The fraction was concentrated under reduced pressure to obtain about 3.8 g of a solid containing MMRC03-0001.
得られた固形物はクロロホルム：メタノール＝１：１の混液４３ｍｌに溶かし、その溶液を５００ｇワコーゲル５０Ｃ１８を１０％アセトニトリル水溶液でつめたカラムに吸着させ、アセトニトリルを段階的に高めていくことにより目的物質のＭＭＲＣ０３−０００１を溶出させた。その結果、ＭＭＲＣ０３−０００１を含有するフラクション０．６Ｌを得た。そのフラクションは減圧下で濃縮後、ＭＭＲＣ０３−０００１を含む約１．６ｇの固形物を得た。得られた固形物はクロロホルム：メタノール＝１：１の混液１０ｍｌに溶かし、その溶液を２００ｇワコーゲル５０Ｃ１８を１０％アセトニトリル水溶液でつめたカラムに吸着させ、流速８ｍｌ／分でアセトニトリルを８時間で１０％から５０％へ直線的に高めることにより目的物質のＭＭＲＣ０３−０００１を溶出させた。その結果、ＭＭＲＣ０３−０００１を含有するフラクション２４０ｍｌを得た。そのフラクションは減圧下で濃縮後、ＭＭＲＣ０３−０００１の５２４ｍｇの無晶形白色粉末を得た。
The obtained solid was dissolved in 43 ml of a mixture of chloroform: methanol = 1: 1, and the solution was adsorbed on a column filled with 500 g Wakogel 50C18 with a 10% acetonitrile aqueous solution. Of MMRC03-0001 was eluted. As a result, a 0.6 L fraction containing MMRC03-0001 was obtained. The fraction was concentrated under reduced pressure to obtain about 1.6 g of a solid containing MMRC03-0001. The obtained solid was dissolved in 10 ml of a mixture of chloroform: methanol = 1: 1, and the solution was adsorbed onto a column filled with 200 g Wakogel 50C18 with a 10% acetonitrile aqueous solution, and acetonitrile was added to 10% in 8 hours at a flow rate of 8 ml / min. The target substance MMRC03-0001 was eluted by linearly increasing from 50 to 50%. As a result, 240 ml of a fraction containing MMRC03-0001 was obtained. The fraction was concentrated under reduced pressure to obtain 524 mg of amorphous white powder of MMRC03-0001.
Claims (8)
１）分子量、分子式：分子量３２６、分子式（Ｃ13Ｈ14Ｎ2Ｏ4Ｓ2）
２）マススペクトル：FAB-MS(NBA)；m/z 349[M+Na]+, 327[M+H]+, 262[M-2S]+
３）紫外線吸収スペクトル：(max nm(() in MeOH; 232 weak shoulder (4270)
４）赤外線吸収スペクトル：(max cm-1 in KBr; 3360, 1680, 1670, 1400, 1360, 1060, 1020, 875, 740, 620
５）核磁気共鳴スペクトル：1H-NMR(400 MHz, CDCl3) ( (ppm): 2.00 (3H, s), 2.83 (1H, m), 3.08 (3H, s), 3.60 (1H, m), 3.65 (1H, m), 4.38 (1H, m), 4.44 (1H, m), 4.63 (1H, m), 5.96 (1H, m)；13C-NMR(100.5 MHz, CDCl3) ( (ppm): 18.3, 27.6, 37.7, 53.7, 55.9, 56.9, 62.2, 73.6, 74.2, 124.2, 134.4, 163.0, 165.8
６）旋光度：[α] D -240( (c=0.38, CHCl3)
７）溶解性：クロロホルム、酢酸エチル、アセトニトリル、アセトン、メタノール、エタノール、ジメチルスルホキシドに易溶、n-ヘキサン、四塩化炭素、水に難溶 Antibiotic MMRC03-0001 having the following physicochemical properties or a pharmacologically acceptable salt thereof:
1) Molecular weight, molecular formula: Molecular weight 326, molecular formula (C 13 H 14 N 2 O 4 S 2)
2) Mass spectrum: FAB-MS (NBA); m / z 349 [M + Na] + , 327 [M + H] + , 262 [M-2S] +
3) UV absorption spectrum: ( max nm (() in MeOH; 232 weak shoulder (4270)
4) Infrared absorption spectrum: ( max cm -1 in KBr; 3360, 1680, 1670, 1400, 1360, 1060, 1020, 875, 740, 620
5) Nuclear magnetic resonance spectrum: 1 H-NMR (400 MHz, CDCl 3 ) ((ppm): 2.00 (3H, s), 2.83 (1H, m), 3.08 (3H, s), 3.60 (1H, m) , 3.65 (1H, m), 4.38 (1H, m), 4.44 (1H, m), 4.63 (1H, m), 5.96 (1H, m); 13 C-NMR (100.5 MHz, CDCl 3 ) ((ppm ): 18.3, 27.6, 37.7, 53.7, 55.9, 56.9, 62.2, 73.6, 74.2, 124.2, 134.4, 163.0, 165.8
6) Optical rotation: [α] D -240 ((c = 0.38, CHCl 3 )
7) Solubility: Easily soluble in chloroform, ethyl acetate, acetonitrile, acetone, methanol, ethanol, dimethyl sulfoxide, hardly soluble in n-hexane, carbon tetrachloride, water
Cepedonium chlorinum TAMA73 strain (FERM P-19617) with the ability to produce the antibiotic MMRC03-0001.
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