JP2573084B2 - Novel antibiotic PF1032B substance and its production method - Google Patents

Novel antibiotic PF1032B substance and its production method

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Publication number
JP2573084B2
JP2573084B2 JP2152862A JP15286290A JP2573084B2 JP 2573084 B2 JP2573084 B2 JP 2573084B2 JP 2152862 A JP2152862 A JP 2152862A JP 15286290 A JP15286290 A JP 15286290A JP 2573084 B2 JP2573084 B2 JP 2573084B2
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Japan
Prior art keywords
pf1032b
substance
antibiotic
culture
methanol
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Other versions
JPH0446188A (en
Inventor
修一 五味
宏臣 渡部
佳代 味戸
千恵子 守山
朋子 湯上
正夫 小山
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Meiji Seika Kaisha Ltd
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Meiji Seika Kaisha Ltd
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Description

【発明の詳細な説明】 産業上の利用分野 本発明は、新規抗生物質PF1032B物質またはその塩、
ならびにその製造法に関する。
The present invention relates to a novel antibiotic PF1032B substance or a salt thereof,
And its manufacturing method.

従来の技術 本発明による抗生物質がPF1032B物質は、上記の理化
学的性状からトリテルペン類あるいはステロイド類と考
えられるが、これと類似する化合物として、本発明者ら
の発見したPF1032物質(特願平1−256980)、ノルトリ
テルペンオリゴ配糖体(特開平1−106897)、エリロシ
ドA(Carmelyら、J.Nat.Prod.52,No.1,167〜170,198
9)等が知られているが、抗生物質PF1032B物質はこれら
の既知物質とは理化学的性状が異なり明確に区別され
る。
2. Description of the Related Art The antibiotic PF1032B according to the present invention is considered to be a triterpene or a steroid from the above-mentioned physicochemical properties. -256980), nortriterpene oligoglycoside (JP-A-1-106689), eryloside A (Carmely et al., J. Nat. Prod. 52, No. 1, 167-170,198).
9) etc., but the antibiotic PF1032B is clearly distinguished from these known substances because of its different physicochemical properties.

発明が解決しようとする課題 従来、微生物が生産する種々の抗生物質が知られてい
るが、毒性の低い抗真菌性抗生物質はそれ程多く見出さ
れていないため、新規な抗真菌性抗生物質の出現が常に
要望されている。本発明者らは、PF1032物質生産菌の培
養物から既知のPF1032物質の他にPF1032B物質が存在す
ることを見出し本発明を単離し、本発明を完成させた。
さらに本発明の目的は、新規抗真菌性抗生物質PF1032B
物質ならびにその製造法を提供することにある。
Problems to be Solved by the Invention Conventionally, various antibiotics produced by microorganisms are known. However, since not many antifungal antibiotics having low toxicity have been found, a new antifungal antibiotic has been developed. Appearance is always desired. The present inventors have found that a PF1032B substance exists in addition to a known PF1032 substance from a culture of a PF1032 substance-producing bacterium, isolated the present invention, and completed the present invention.
It is a further object of the present invention to provide a novel antifungal antibiotic PF1032B
An object of the present invention is to provide a substance and a method for producing the substance.

課題を解決するための手段 第1の本発明の要旨とするところは、新規抗生物質PF
1032B物質およびその塩にある。本発明によるPF1032B物
質の理化学的および生物学的性状は、次の通りである。
Means for Solving the Problems The gist of the first invention is to provide a novel antibiotic PF.
1032B substance and its salts. The physicochemical and biological properties of the PF1032B substance according to the present invention are as follows.

1.PF1032B物質の理化学的性状 (1) 色および形状:白色粉末 (2) 元素分析:C35H54O8・H2Oとして 実測値 C 67.78%、H 9.20% 計算値 C 67.71%、H 9.09% (3) マススペクトル(FD−MS):m/z602(M+) (4) 融点:192〜195℃(分解) (5) 比旋光度:▲〔α〕22 D▼=+106.7゜(cl.0,M
eOH) (6) 紫外部吸収スペクトル 〔MeOH〕:206(125),226(sh 44) 〔0.1N HCl−MeOH〕:205(113),227(sh 39) 〔0.1N NaOH−MeOH〕:214(198) (7) 赤外部吸収スペクトル (KBr cm-1):3400,2920,2870,1690,1650,1460,1450,14
40,1405,1380,1245,1200,1155,1115,1060,1040,910,86
0,785,720 (8) 1H NMRスペクトル(400MHz、CD3OD) δ(ppm):0.81(3H,s),0.91(3H,s),0.94(3H,d),
1.05(1H,m),1.35〜1.52(5H,m),1.52〜1.76(5H,
m),1.59(3H,br s),1.66(3H,br s),1.88(2H,m),
1.94(1H,br dd),1.97〜2.19(7H,m),2.26(1H,m),
2.29(1H,m),3.30(1H,dd),3.41(1H,dd),3.63(1H,
m),3.65(1H,m),3.73(1H,m),3.76(1H,dd),4.09
(1H,br dd),4.66(1H,br s),5.00(1H,d),5.08(1
H,br dd),5.26(1H,br s) (9) 13C NMRスペクトル(100MHz,CD3OD) δ(ppm):179.8 s,151.3 s,140.1 s,131.8 s,129.1 s,
126.1 d,104.7 t,96.9 d,76.7 d,75.2 d,74.0 d,73.6
d,71.9 d,63.8 s,62.7 t,52.1 d,48.1 s,47.8 d,40.4
s,37.2 t,37.0 d,36.1 t,32.7 t,30.2 t,29.4 t,29.0
t,27.7 t,25.9 q,25.7 t,24.4 t,22.0 t,19.1 q,18.6
q,18.3 q,17.1 q (10) 溶解性:メタノール、アセトン、酢酸エチルに
溶け、クロロホルムに溶け難く、水に極めて溶け難い。
1. Physical and chemical properties of PF1032B substance (1) Color and shape: white powder (2) Elemental analysis: C 35 H 54 O 8 · H 2 O Measured value C 67.78%, H 9.20% Calculated value C 67.71%, H 9.09% (3) Mass spectrum (FD-MS): m / z 602 (M + ) (4) Melting point: 192 to 195 ° C. (decomposition) (5) Specific rotation: ▲ [α] 22 D ▼ = +106.7゜ (cl.0, M
eOH) (6) Ultraviolet absorption spectrum [MeOH]: 206 (125), 226 (sh44) [0.1N HCl-MeOH]: 205 (113), 227 (sh39) [0.1N NaOH-MeOH]: 214 (198) (7) Infrared absorption Spectrum (KBr cm -1 ): 3400,2920,2870,1690,1650,1460,1450,14
40,1405,1380,1245,1200,1155,1115,1060,1040,910,86
0,785,720 (8) 1 H NMR spectrum (400 MHz, CD 3 OD) δ (ppm): 0.81 (3H, s), 0.91 (3H, s), 0.94 (3H, d),
1.05 (1H, m), 1.35 to 1.52 (5H, m), 1.52 to 1.76 (5H,
m), 1.59 (3H, br s), 1.66 (3H, br s), 1.88 (2H, m),
1.94 (1H, br dd), 1.97 ~ 2.19 (7H, m), 2.26 (1H, m),
2.29 (1H, m), 3.30 (1H, dd), 3.41 (1H, dd), 3.63 (1H,
m), 3.65 (1H, m), 3.73 (1H, m), 3.76 (1H, dd), 4.09
(1H, br dd), 4.66 (1H, br s), 5.00 (1H, d), 5.08 (1
H, br dd), 5.26 (1H, br s) (9) 13 C NMR spectrum (100 MHz, CD 3 OD) δ (ppm): 179.8 s, 151.3 s, 140.1 s, 131.8 s, 129.1 s,
126.1 d, 104.7 t, 96.9 d, 76.7 d, 75.2 d, 74.0 d, 73.6
d, 71.9 d, 63.8 s, 62.7 t, 52.1 d, 48.1 s, 47.8 d, 40.4
s, 37.2 t, 37.0 d, 36.1 t, 32.7 t, 30.2 t, 29.4 t, 29.0
t, 27.7 t, 25.9 q, 25.7 t, 24.4 t, 22.0 t, 19.1 q, 18.6
q, 18.3 q, 17.1 q (10) Solubility: Soluble in methanol, acetone, ethyl acetate, hardly soluble in chloroform, and extremely hardly soluble in water.

(11) 塩基性、酸性、中性の区別:弱酸性物質 PF1032B物質の塩としては、金属塩、特にナトリウム
塩の如きアルカリ金属塩、カルシウム塩の如きアルカリ
土類金属塩があり、更に後記の如きアミンとの塩等があ
る。
(11) Basic, acidic and neutral distinctions: Weakly acidic substances Salts of the PF1032B substance include metal salts, particularly alkali metal salts such as sodium salts and alkaline earth metal salts such as calcium salts. Such as salts with amines.

2.PF1032物質の生物活性 本発明によるPF1032B物質の各種真菌および各種細菌
に対する最小発育阻止濃度をそれぞれ第1表および第2
表に示した。
2. Biological activity of PF1032 substance The minimum inhibitory concentrations of the PF1032B substance of the present invention against various fungi and various bacteria are shown in Tables 1 and 2, respectively.
It is shown in the table.

3.PF1032B物質の急性毒性 PF1032B物質のJcl:ICRマウスおよび(雄、19〜20g、
n=3)を用いた腹控内投与による急性毒性試験で、10
0gm/kgの投与量で死亡例はなかった。
3. Acute toxicity of PF1032B substance Jcl of PF1032B substance: ICR mice and (male, 19-20 g,
In an acute toxicity test by intra-abdominal administration using n = 3), 10
No deaths occurred at the dose of 0 gm / kg.

第2の本発明の要旨とするところは、不整子のう菌に
属する抗生物質PF1032B物質生産菌を培養し、その培養
物から抗生物質PF1032B物質を採取する抗生物質PF1032B
物質の製造法にある。
According to a second aspect of the present invention, an antibiotic PF1032B is produced by culturing an antibiotic PF1032B substance-producing bacterium belonging to the arrhythmic bacteria and collecting the antibiotic PF1032B substance from the culture.
In the method of manufacturing the substance.

本発明の抗生物質PF1032B物質を生産するために使用
される微生物の実用的な例としては、ネパールの土壌よ
り分離したネオサルトリヤ属に属するPF1032株が挙げら
れる。
Practical examples of microorganisms used for producing the antibiotic PF1032B of the present invention include the strain PF1032 belonging to the genus Neosartria isolated from soil in Nepal.

1.PF1032株の菌学的性状 I.各培地における生育状態 (1) ツアペック寒天培地 生育はあまり良くなく25℃では培養7日間で直径が8
〜10mm、14日間で22〜25mm、37℃では培養7日間で直径
が18〜20mm、14日間で30〜35mmである。白色、ビロード
状の菌糸からなり、子のう果、分生子構造は殆ど形成さ
れない。
1. Mycological properties of strain PF1032 I. Growth state in each medium (1) Tuapek agar medium Growth is not so good, and the diameter is 8 in 7 days of culture at 25 ° C.
1010 mm, 22 to 25 mm for 14 days, 18 to 20 mm in diameter for 7 days at 37 ° C., 30 to 35 mm for 14 days. Consisting of white, velvety hyphae, almost no ascocarp and conidia are formed.

(2) ツアペック酵母エキス寒天培地 37℃で生育が速く、培養7日間で直径が70〜80mmに達
する。平坦な集落となり、子のう果は層状に密生し、最
初は白色であるが後に淡灰茶色となる。分生子構造はあ
まり形成しない。集落裏面は淡黄褐色となる。25℃にお
いても37℃と同様の性状であるが、生育速度は培養7日
間で直径が55〜60mmとやや遅く、子のう果の形成も少し
悪くなる。
(2) Tuapec yeast extract agar medium The growth is fast at 37 ° C., and the diameter reaches 70 to 80 mm in 7 days of culture. It becomes a flat settlement, and the ascocarps grow densely in layers, initially white, but later pale gray brown. Conidia structures do not form much. The backside of the settlement becomes pale yellow-brown. At 25 ° C., the properties are the same as at 37 ° C., but the growth rate is slightly slower at 55 to 60 mm in diameter during 7 days of culture, and the formation of ascocarp is slightly worse.

(3) 麦芽エキス寒天培地 37℃で生育が速く、培養7日間で直径が85mm以上に達
する。性状はツアペック酵母エキス寒天培地とほぼ同様
であるが、分生子構造をよく形成する。
(3) Malt extract agar medium It grows rapidly at 37 ° C and reaches a diameter of 85 mm or more in 7 days of culture. The properties are almost the same as those of Tuapec yeast extract agar medium, but they form conidia well.

II.顕微鏡下における形態的特色 (1) 閉子のう殻 表在性、白色〜乳白色、球形〜亜球形、直径は90〜15
0μm、不稔性菌糸で緩く覆われる。
II. Morphological features under microscope (1) Closed cysts Superficial, white to milky white, spherical to subspherical, 90 to 15 in diameter
0 μm, loosely covered with sterile hyphae.

(2) 子のう 8胞子、無色、球形〜亜球形、直径は13〜16μm、消
失性。
(2) Ascosac 8 spores, colorless, spherical to subspherical, 13 to 16 μm in diameter, vanishing.

(3) 子のう胞子 無色、レンズ形、大きさは3.6〜4.0×5.0〜6.0μm、
2本の赤道面隆起を生じ、光学顕微鏡のレベルでは殆ど
滑面である。
(3) Ascospores colorless, lens shape, size is 3.6 ~ 4.0 × 5.0 ~ 6.0μm,
It produces two equatorial ridges and is almost smooth at the level of an optical microscope.

(4) 分生子頭 ゆるい円筒形、白色。(4) Conidium head Loose cylindrical, white.

(5) フィアライド 頂のうの上部1/2より生じ単列。(5) Fialide A single row from the upper half of the crown.

(6) 分生子 球形〜亜球形、直径は2.0〜2.4μm。(6) Conidia spherical to subspherical, 2.0 to 2.4 μm in diameter.

本菌の菌学的性状は以上の通りであるが、閉子のう殻
を形成し、分生子世代はアスペルギルス(Aspergillu
s)属であることから、不整子のう菌綱コウジカビ科に
属する。さらに閉子のう殻が白色〜乳白色であり、不稔
性菌糸に緩く覆われ、分生子世代はアスペルギルス・フ
ミガタス(Aspergillus fumigatus)菌群であることか
らネオサルトリア(Neosartorya)属に属するものと考
えられる。
The mycological properties of this bacterium are as described above, but it forms a closed cyst and the conidium generation is Aspergillus (Aspergillus).
s) Because it belongs to the genus, it belongs to the class of Aspergillus oryzae. Furthermore, it is considered that it belongs to the genus Neosartorya (Aspergillus fumigatus) because it has white to milky white husks, is loosely covered with sterile hyphae, and has a conidium generation of Aspergillus fumigatus. Can be

従って、本菌株を不整子のう菌PF1032株と呼称するこ
とにした。尚、本菌株は工業技術院微生物工業技術研究
所に微工研菌寄第11036号(FERM P−11036)として寄託
されている。
Therefore, this strain was referred to as the arachnoid fungus strain PF1032. This strain has been deposited with the Institute of Microbial Industry and Technology of the National Institute of Advanced Industrial Science and Technology as Microbial Research Bacteria No. 11036 (FERM P-11036).

PF1032株は、他のカビに見られるようにその性状が変
化し安い。例えば、PF1032株に由来する突然変異株(自
然発生または誘発性)、形質接合体また遺伝子組換え体
であっても、PF1032B物質を生産するものは全て本発明
に使用できる。
The PF1032 strain changes its properties and is cheap as seen in other molds. For example, any mutants (naturally occurring or inducible) derived from the PF1032 strain, transgenes, or transgenics that produce the PF1032B substance can be used in the present invention.

2.PF1032B物質生産菌の培養法 不整子のう菌に属するPF1032B物質生産菌を通常の微
生物が利用しうる栄養物を含有する培地で培養する。栄
養源としては、従来カビの培養に利用されている公知の
ものが使用できる。例えば、炭酸源としては、グルコー
ス、シュクロース、水飴、デキストリン、澱粉、グリセ
ロール、糖蜜、動・植物油等を使用しうる。また、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、ペプトン、酵母エキス、硫
酸アンモニウム、硝酸ナトリウム、尿素等を使用しう
る。その他必要に応じ、ナトリウム、カリウム、カルシ
ウム、マグネシウム、コバルト、塩素、燐酸、硫酸およ
びその他のイオンを生成することができる無機塩類を添
加することは有効である。また、菌の発育を助け、PF10
32B物質の生産を促進するような有機および無機物を適
当に添加することができる。
2. Culture method of PF1032B substance producing bacterium The PF1032B substance producing bacterium belonging to the arbuscular bacillus is cultured in a medium containing nutrients that can be used by ordinary microorganisms. As nutrient sources, known nutrients conventionally used for mold culture can be used. For example, glucose, sucrose, starch syrup, dextrin, starch, glycerol, molasses, animal and vegetable oils and the like can be used as the carbonic acid source. Nitrogen sources include soy flour, wheat germ, corn steep,
Liquor, cottonseed meal, meat extract, peptone, yeast extract, ammonium sulfate, sodium nitrate, urea and the like can be used. If necessary, it is effective to add sodium, potassium, calcium, magnesium, cobalt, chlorine, phosphoric acid, sulfuric acid and other inorganic salts capable of producing ions. It also helps the growth of fungi, PF10
Organic and inorganic substances that promote the production of the 32B substance can be suitably added.

培養法としては、好気的条件での培養法、特に深部培
養法が最も適している。培養に適当な温度は15〜40℃で
あるが、多くの場合26℃付近で培養する。PF1032B物質
の生産は培地や培養条件により異なるが、振盪培養、タ
ンク培養のいずれにおいても通常2〜10日間でその蓄積
が最高に達する。培養中のPF1032B物質の蓄積量が最高
になった時に培養を停止し、培養液から目的物質を単離
精製する。
As a culture method, a culture method under aerobic conditions, particularly a submerged culture method is most suitable. A suitable temperature for culturing is 15 to 40 ° C, but the culturing is often performed at around 26 ° C. The production of the PF1032B substance varies depending on the culture medium and culture conditions, but usually reaches the maximum in both shaking culture and tank culture in 2 to 10 days. When the accumulated amount of the PF1032B substance in the culture reaches the maximum, the culture is stopped, and the target substance is isolated and purified from the culture solution.

3.PF1032B物質の精製法 本発明によって得られるPF1032B物質の培養物からの
採取に当たっては、その性状を利用した通常の分離手
段、例えば、溶剤抽出法、イオン交換樹脂法、吸着また
は分配カラムクロマト法、ゲルろ過法、透析法、沈澱法
等を単独でまたは適宜組み合わせて抽出精製することが
できる。例えば、PF1032B物質は、培養菌体中からはア
セトン−水、メタノール−水または酢酸エチル等で抽出
される。また、培養液中に蓄積されたPF1032B物質は、
水と混ざらない有機溶剤、例えば、ブタノール、酢酸エ
チル等で抽出すればPF1032B物質は有機溶剤層に抽出さ
れる。
3.Purification method of PF1032B substance In collecting the PF1032B substance obtained by the present invention from a culture, a usual separation method utilizing its properties, for example, a solvent extraction method, an ion exchange resin method, an adsorption or distribution column chromatography method , Gel filtration, dialysis, precipitation, etc., alone or in combination as appropriate. For example, the PF1032B substance is extracted from the cultured cells with acetone-water, methanol-water, ethyl acetate, or the like. Also, the PF1032B substance accumulated in the culture solution is
Extraction with an organic solvent immiscible with water, for example, butanol, ethyl acetate, etc., will extract the PF1032B substance into the organic solvent layer.

PF1032B物質を更に精製するには、シリカゲル(ワコ
ーゲル C−300、和光純薬工業社製等)、アルミナ等
の吸着剤やセファデックスLH−20(ファルマシア社
製)、トヨパール HW−40(株式会社東ソー社製)等を
用いるクロマトグラフィーを行うとよい。
To further purify the PF1032B substance, use silica gel (Wakogel C-300, manufactured by Wako Pure Chemical Industries, Ltd.), adsorbent such as alumina, Sephadex LH-20 (manufactured by Pharmacia), Toyopearl HW-40 (Tosoh Corporation) It is preferable to carry out chromatography using, for example, the following method.

このようにして培養物中に生産されたPF1032B物質は
遊離の形、すなわちPF1032B物質それ自体として分離す
ることができ、またPF1032B物質を含有する溶液または
その濃縮液を塩基、すなわち例えば水酸化ナトリウム、
水酸化カリウム等のアルカリ金属化合物、水酸化カルシ
ウム、水酸化マグネシウム等のアルカリ土類金属化合
物、アンモニウム塩等のような無機塩基、エタノールア
ミン、トリエチルアミン、ジシクロヘキシルアミン等の
有機塩基により、各工程の操作中例えば抽出、分離また
は精製の各工程の操作中に処理した場合、PF1032B物質
は対応するその塩類の形に変化し分離される。また別に
このようにして製造されたPF1032B物質の塩類は、常法
により遊離の形、PF1032B物質それ自体に変化させるこ
とができる。更に遊離の形で得られたPF1032B物質を前
記塩基により常法で対応するその塩類に変化させてもよ
い。従ってPF1032B物質と同様に前記のようなその塩類
も、この発明の範囲内に包含されるものとする。
The PF1032B substance thus produced in the culture can be separated in free form, i.e., as the PF1032B substance itself, and the solution containing the PF1032B substance or its concentrated solution can be treated with a base, i.e. sodium hydroxide, for example.
Operation of each step using an alkali metal compound such as potassium hydroxide, an alkaline earth metal compound such as calcium hydroxide or magnesium hydroxide, an inorganic base such as an ammonium salt, or an organic base such as ethanolamine, triethylamine or dicyclohexylamine. During processing, for example during the operation of each step of extraction, separation or purification, the PF1032B substance changes to its corresponding salt form and is separated. Alternatively, the salts of the PF1032B substance thus produced can be converted into a free form, the PF1032B substance itself, by a conventional method. Further, the PF1032B substance obtained in the free form may be converted into the corresponding salts by a conventional method using the base. Accordingly, salts thereof as described above, as well as the PF1032B material, are intended to be included within the scope of the present invention.

実施例 以下に本発明の実施例を示すが、PF1032B物質の性状
が本発明によって明らかにされたので、それらの性状に
もとづきPF1032B物質の製造法を種々考案することがで
きる。従って本発明は実施例に限定されるものではな
く、実施例の修飾手段は勿論、本発明によって明らかに
されたPF1032B物質の性状にもとづいて公知の手段を施
してPF1032B物質を生産、濃縮、抽出、精製する方法を
すべて包括する。
Examples Examples of the present invention will be described below. Since the properties of the PF1032B substance have been clarified by the present invention, various methods for producing the PF1032B substance can be devised based on those properties. Therefore, the present invention is not limited to the examples, and it is possible to produce, concentrate, and extract the PF1032B substance by applying known means based on the properties of the PF1032B substance revealed by the present invention, as well as the modification means of the examples. , Encompasses all purification methods.

種培地として、スターチ2.0%、グルコース1.0%、ポ
リペプトン0.5%、小麦胚芽0.6%、酵母エキス0.3%、
大豆粕0.2%、炭酸カルシウム0.2%の組成からなる培地
を用いた。また、生産培地として、水飴2.0%、大豆油
0.3%、大豆粕1.2%、小麦胚芽1.2%、硫酸ナトリウム
0.02%、硫酸鉄(7水塩)0.0005%、塩化コバルト(7
水塩)0.0005%、炭酸カルシウム0.1%の組成からなる
培地を用いた。なお、殺菌前pHはすべてpH7.0に調整し
て使用した。
As a seed medium, starch 2.0%, glucose 1.0%, polypeptone 0.5%, wheat germ 0.6%, yeast extract 0.3%,
A medium having a composition of soybean meal 0.2% and calcium carbonate 0.2% was used. In addition, as a production medium, 2.0% syrup, soybean oil
0.3%, soybean meal 1.2%, wheat germ 1.2%, sodium sulfate
0.02%, iron sulfate (heptahydrate) 0.0005%, cobalt chloride (7
A medium having a composition of (water salt) 0.0005% and calcium carbonate 0.1% was used. The pH before sterilization was all adjusted to pH 7.0 before use.

前記の種培地(20ml)を分注した100ml容三角フラス
コを120℃で15分間殺菌し、これにNeosartorya属PF1032
株(FERM P−11036)の斜面寒天培養の1白金耳を接種
し、26℃で2日間振盪培養して種培養とした。次いで、
前記の生産培地(100ml)を分注した500ml容三角フラス
コ(200本)を120℃で15分間殺菌し、これに前記種培養
(各2ml)を接種し、26℃で4日間振盪培養した。培養
終了後、濾過助剤として珪藻土を加えて濾過し、濾液と
菌体を得た。
A 100 ml Erlenmeyer flask to which the above seed medium (20 ml) was dispensed was sterilized at 120 ° C. for 15 minutes, and Neosartorya sp.
One platinum loop of the slant agar culture of the strain (FERM P-11036) was inoculated and shake-cultured at 26 ° C for 2 days to obtain a seed culture. Then
A 500-ml Erlenmeyer flask (200 tubes) into which the production medium (100 ml) was dispensed was sterilized at 120 ° C. for 15 minutes, inoculated with the seed culture (2 ml each), and cultured with shaking at 26 ° C. for 4 days. After the completion of the culture, diatomaceous earth was added as a filter aid, followed by filtration to obtain a filtrate and cells.

この菌体に50%アセトン水(16L)を加え、1時間撹
拌後菌体を濾別して菌体抽出液を得た。菌体抽出液は、
減圧下でアセトンを留去して4Lの濃縮液とした。この濃
縮液と培養濾液(16L)の混合液から酢酸エチル(20L)
で活性成分を抽出し、酢酸エチル層を濃縮すると油状物
質(3.82g)が得られた。この油状物質をシリカゲルカ
ラム(400g)の上部に載せ、クロロホルム−メタノール
(10:1,800ml)で洗浄後クロロホルム−メタノール(5:
1)を展開溶媒とするクロマトグラフィーを行い、溶出
液を20mlずつ分画した。PF1032B物質を含む画分(フラ
クション番号81〜90)を濃縮乾固すると淡黄色粉末(3
2.3mg)が得られた。次いでこの淡黄色粉末を、メタノ
ールを展開溶媒とするセファデックスLH−20(140ml)
カラムクロマトグラフィー(1分画,6.5ml)を行って精
製し、フラクション番号15〜19を濃縮乾固すると純粋な
PF1032B物質が白色粉末(25.8mg)として得られた。同
様に、最初のシリカゲル・カラムクロマトグラフィーに
於いてPF1032物質を含む画分(フラクション番号92〜13
6)を濃縮乾固すると淡黄色粉末(214mg)が得られた。
次いで、この淡黄色粉末をメタノールを展開溶媒とする
セファデックスLH−20(560ml)カラムクロマトグラフ
ィー(1分画,13ml)を行って精製し、フラクション番
号34〜42を濃縮乾固すると純粋なPF1032物質が白色粉末
(189mg)として得られた。
50% acetone water (16 L) was added to the cells, and after stirring for 1 hour, the cells were filtered off to obtain a cell extract. The cell extract is
Acetone was distilled off under reduced pressure to obtain a 4 L concentrated solution. Ethyl acetate (20L) was obtained from the mixture of this concentrated solution and culture filtrate (16L).
The ethyl acetate layer was concentrated to give an oily substance (3.82 g). This oily substance was placed on the top of a silica gel column (400 g), washed with chloroform-methanol (10: 1, 800 ml), and then chloroform-methanol (5: 8).
Chromatography was performed using 1) as a developing solvent, and the eluate was fractionated in 20 ml portions. The fraction containing the PF1032B substance (fraction numbers 81 to 90) was concentrated to dryness to give a pale yellow powder (3
2.3 mg) were obtained. Next, this pale yellow powder was separated with Sephadex LH-20 (140 ml) using methanol as a developing solvent.
Purification was performed by column chromatography (1 fraction, 6.5 ml), and fractions 15 to 19 were concentrated to dryness to give pure
The PF1032B material was obtained as a white powder (25.8mg). Similarly, in the first silica gel column chromatography, the fraction containing the PF1032 substance (fraction numbers 92 to 13)
6) was concentrated to dryness to give a pale yellow powder (214 mg).
Then, the pale yellow powder was purified by column chromatography (1 fraction, 13 ml) using Sephadex LH-20 (560 ml) using methanol as a developing solvent, and fractions 34 to 42 were concentrated to dryness to give pure PF1032. Material was obtained as a white powder (189 mg).

発明の効果 本発明のPF1032B物質は第1表に示した如く抗真菌活
性を有し、また従来の抗真菌性抗生物質に比べて毒性が
低いのが特徴である。これらの性質に基づき本発明のPF
1032B物質を抗真菌剤あるいはそれへの変換用素材とし
て用いることができる。
Effect of the Invention The PF1032B substance of the present invention is characterized by having antifungal activity as shown in Table 1 and having lower toxicity than conventional antifungal antibiotics. Based on these properties, the PF of the present invention
The 1032B substance can be used as an antifungal agent or a material for conversion to it.

【図面の簡単な説明】[Brief description of the drawings]

第1図:PF1032B物質のメタノール中(100μg/ml,実
線)、酸性メタノール中(100μg/ml,破線)および塩基
性メタノール中(100μg/ml,一点鎖線)での紫外部吸収
スペクトルを示す。 第2図:PF1032B物質の臭化カリウム錠での赤外部吸収ス
ペクトルを示す。 第3図:PF1032B物質の重メタノール溶液中での400MHz 1
H NMRスペクトルを示す。 第4図:PF1032B物質の重メタノール溶液中での100MHz
13C NMRスペクトルを示す。
FIG. 1: UV absorption spectra of the PF1032B substance in methanol (100 μg / ml, solid line), acidic methanol (100 μg / ml, broken line) and basic methanol (100 μg / ml, dashed line). FIG. 2 shows an infrared absorption spectrum of a potassium bromide tablet of the PF1032B substance. Figure 3: PF1032B 400MHz 1 in heavy methanol solution of a substance
1 shows a 1 H NMR spectrum. Figure 4: 100MHz in PF1032B substance in heavy methanol solution
13 shows a 13 C NMR spectrum.

フロントページの続き (72)発明者 守山 千恵子 神奈川県横浜市港北区師岡町760 明治 製菓株式会社薬品総合研究所内 (72)発明者 湯上 朋子 神奈川県横浜市港北区師岡町760 明治 製菓株式会社薬品総合研究所内 (72)発明者 小山 正夫 神奈川県横浜市港北区師岡町760 明治 製菓株式会社薬品総合研究所内Continued on the front page (72) Inventor Chieko Moriyama 760, Shiokaokacho, Kohoku-ku, Yokohama, Kanagawa Prefecture Inside the Pharmaceutical Research Institute (72) Inventor Tomoko Yugami 760, Shikaokacho, Kohoku-ku, Yokohama-shi, Kanagawa Meiji Seika Co., Ltd. Inside the Research Institute (72) Inventor Masao Koyama 760 Meijiokacho, Kohoku-ku, Yokohama-shi, Kanagawa Prefecture Meiji Seika Co., Ltd.

Claims (2)

(57)【特許請求の範囲】(57) [Claims] 【請求項1】下記の特性を有する抗生物質PF1032B物質
およびその塩 (1) 色および形状:白色粉末 (2) 元素分析:実測値 C 67.78%,H 9.20% (3) マススペクトル(FD−MS):m/z602 (4) 融点:192〜195℃(分解) (5) 比旋光度:▲〔α〕22 D▼=+106.7゜(cl.0,M
eOH) (6) 紫外部吸収スペクトル:第1図に示す。 (7) 赤外部吸収スペクトル:第2図に示す。 (8) 1H NMRスペクトル:第3図に示す。 (9) 13C NMRスペクトル:第4図に示す。 (10) 溶解性:メタノール、アセトン、酢酸エチルに
溶け、クロロホルムに溶け難く、水に極めて溶け難い。 (11) 塩基性、酸性、中性の区別:逆酸性物質
1. Antibiotic PF1032B substance and its salt having the following properties: (1) Color and shape: white powder (2) Elemental analysis: measured values C 67.78%, H 9.20% (3) Mass spectrum (FD-MS) ): M / z602 (4) Melting point: 192 to 195 ° C (decomposition) (5) Specific rotation: ▲ [α] 22 D ▼ = +106.7 ゜ (cl.0, M
eOH) (6) Ultraviolet absorption spectrum: shown in FIG. (7) Infrared external absorption spectrum: shown in FIG. (8) 1 H NMR spectrum: shown in FIG. (9) 13 C NMR spectrum: shown in FIG. (10) Solubility: soluble in methanol, acetone, ethyl acetate, hardly soluble in chloroform, and extremely hardly soluble in water. (11) Basic, acidic, and neutral: reverse acidic substances
【請求項2】不整子のう菌(Neosartorya属)に属す
る、抗生物質PF1032B物質生産菌を培養し、その培養物
から抗生物質PF1032B物質を採取することを特徴とする
抗生物質PF1032B物質の製造法。
2. A method for producing an antibiotic PF1032B, comprising culturing an antibiotic PF1032B substance-producing bacterium belonging to the arachnoid bacterium (genus Neosartorya) and collecting the antibiotic PF1032B from the culture. .
JP2152862A 1990-06-13 1990-06-13 Novel antibiotic PF1032B substance and its production method Expired - Lifetime JP2573084B2 (en)

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JP2152862A JP2573084B2 (en) 1990-06-13 1990-06-13 Novel antibiotic PF1032B substance and its production method

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Application Number Priority Date Filing Date Title
JP2152862A JP2573084B2 (en) 1990-06-13 1990-06-13 Novel antibiotic PF1032B substance and its production method

Publications (2)

Publication Number Publication Date
JPH0446188A JPH0446188A (en) 1992-02-17
JP2573084B2 true JP2573084B2 (en) 1997-01-16

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Country Link
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* Cited by examiner, † Cited by third party
Publication number Priority date Publication date Assignee Title
US5521169A (en) * 1994-01-14 1996-05-28 Bristol-Myers Squibb Company Ascosteroside and analogs thereof useful in antifungal compositions for methods of treating infections and inhibition of fungal growth
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