JPS62135497A - P4 substance and preparation thereof - Google Patents
P4 substance and preparation thereofInfo
- Publication number
- JPS62135497A JPS62135497A JP60275220A JP27522085A JPS62135497A JP S62135497 A JPS62135497 A JP S62135497A JP 60275220 A JP60275220 A JP 60275220A JP 27522085 A JP27522085 A JP 27522085A JP S62135497 A JPS62135497 A JP S62135497A
- Authority
- JP
- Japan
- Prior art keywords
- substance
- water
- ethanol
- methanol
- bacillus
- Prior art date
- Legal status (The legal status is an assumption and is not a legal conclusion. Google has not performed a legal analysis and makes no representation as to the accuracy of the status listed.)
- Pending
Links
Abstract
Description
【発明の詳細な説明】
本発明は、新規なＰ４物質及びその′Ａ造法に関するも
のである。DETAILED DESCRIPTION OF THE INVENTION The present invention relates to a novel P4 substance and its 'A production method.
更に詳細には、本発明は、アミノ酸９分子と未知酸１分
子からなる新規な抗真菌性Ｐ４物質及びその製造法に関
するものである。More specifically, the present invention relates to a novel antifungal P4 substance consisting of nine amino acid molecules and one unknown acid molecule, and a method for producing the same.
本発明者らは、バチルス属の生産する低分子抗菌物質を
求めて鋭意研究した結果、バチルス・ズブチリスの培養
物から新規な抗真菌性物質を単離することに成功したの
である。新たに単離された抗真菌性物質はＰ４物質と命
名された。As a result of intensive research in search of low-molecular antibacterial substances produced by Bacillus, the present inventors succeeded in isolating a novel antifungal substance from a culture of Bacillus subtilis. The newly isolated antifungal substance was named P4 substance.
Ｐ４物質の生産菌としては、Ｐ４物質を生産す゛ればい
かなる菌でもよいが、バチルス・ズブチリスＮｏ、０６
０．ＦＥＲＭ Ｐ−８５４２を用いるのが好ましい。Any bacteria that produces P4 substances may be used as the P4 substance producing bacteria, but Bacillus subtilis No. 06
0. Preferably, FERM P-8542 is used.
Ｐ４物質生産菌の培養培地は、資化できる炭素源、窒素
源、ビタミン等を含有する栄養料等適宜含有したもので
、Ｐ４物質を生産蓄積する培地であればいずれの培地で
もよい。The culture medium for the P4 substance-producing bacteria may contain appropriate nutrients such as assimilable carbon sources, nitrogen sources, vitamins, etc., and may be any medium that can produce and accumulate P4 substances.
バチルス・ズブチリスＮｏ、０６０の培養に適した培地
は例えば次のものがあげられる。Examples of suitable media for culturing Bacillus subtilis No. 060 include the following.
ポリペプトン ３０ｇ
酵母エキス ５ｇ
ＮａＣ０５ｇ
脱イオン水 ＩＱ
（ｐＨ７，０）
培養は２５〜３０℃で１通気、攪拌、振どう等好気的に
４日間程度行なわれる。Polypeptone 30g Yeast extract 5g NaC 05g Deionized water IQ (pH 7,0) Cultivation is carried out aerobically at 25-30°C for about 4 days with ventilation, stirring, shaking, etc.
得られた培養液は濾過して、濾液を得、これに６Ｎ ｔ
ｌｃＩ２を添加し、ρ１（＝３．０に調整し沈澱物を得
る。The obtained culture solution was filtered to obtain a filtrate, and 6N t was added to this.
Add lcI2 and adjust to ρ1 (=3.0) to obtain a precipitate.
沈澱物は水に溶解し、ＮａＨＣＯ，を加え、ｐＨ＝７．
０とし１次いでエタノールを８０％まで加える。得られ
た上清液を真空乾燥し、乾燥物を蒸留水に溶解し、再び
６Ｎ ＨＣＱを加えてｐＨ＝３．０に調整し、沈澱物を
得る。得られた沈澱物を蒸留水に溶解し、ＮａｔｌＣＯ
。The precipitate was dissolved in water and NaHCO was added to pH=7.
0 and then add ethanol to 80%. The obtained supernatant liquid is vacuum-dried, the dried product is dissolved in distilled water, and 6N HCQ is added again to adjust the pH to 3.0 to obtain a precipitate. The obtained precipitate was dissolved in distilled water and NatlCO
.
を加え、ｐＨ＝７．０とし、蒸留水に対して透析する。was added to adjust the pH to 7.0, and the mixture was dialyzed against distilled water.
透析内液を凍結乾燥することによってＰ４物質含有粗精
製物を黄色粉末として得る。By freeze-drying the dialyzed fluid, a crude product containing P4 substance is obtained as a yellow powder.
得られた黄色粉末を水に溶解し、アセトニトリル系で高
速液体クロマトにかけ、これを単離し、凍結乾燥するこ
とによってほぼ純品のＰ４物質を白色無定形粉末として
得る。The obtained yellow powder is dissolved in water, subjected to high performance liquid chromatography using acetonitrile system, isolated, and freeze-dried to obtain substantially pure P4 substance as a white amorphous powder.
得られたＰ４物質の理化学的性質は次の通りである。The physicochemical properties of the obtained P4 substance are as follows.
１、紫外線吸収スペクトルは第１図に示す通りである。1. The ultraviolet absorption spectrum is as shown in FIG.
２、溶解性：エタノール、メタノール、アセトン、水に
不溶で、希エタノール水、希メタノール水、ＣＨＣＱ３
に可溶である。2. Solubility: Insoluble in ethanol, methanol, acetone, water, dilute ethanol water, dilute methanol water, CHCQ3
It is soluble in
３、物質の色、性状：白色無定形粉末
４、呈色反応：ニンヒドリンテスト
５、一本物質のアミノ酸組成：本物質の加水分解物のア
ミノ酸分析によって次の結果を得た。3. Color and properties of the substance: White amorphous powder 4. Color reaction: Ninhydrin test 5. Amino acid composition of the substance: The following results were obtained by amino acid analysis of the hydrolyzate of this substance.
６、本物質は真菌に対して強い抗菌性を示す。6. This substance exhibits strong antibacterial properties against fungi.
サッカロミセス・セレビシエＩＡＭ ４１２５の生育を
１６０μｇ／１２で阻止する。Growth of Saccharomyces cerevisiae IAM 4125 is inhibited at 160 μg/12.
７、精製法：ｐＨ＝３．０程度で沈澱させ、沈澱物をｐ
Ｈ＝７．０で溶解する工程をくりかえし、最後に高速液
体クロマトにかけることによってほぼ純品を得ることが
できる。高速液体クロマトにおける入２１０による吸光
度曲線は第２図に示す通りである。矢印がＰ４物質を示
している。7. Purification method: Precipitate at pH = about 3.0, and remove the precipitate from pH
By repeating the dissolution step at H=7.0 and finally applying it to high performance liquid chromatography, an almost pure product can be obtained. The absorbance curve obtained by input 210 in high performance liquid chromatography is as shown in FIG. The arrow indicates the P4 substance.
次に本発明の実施例を示す。Next, examples of the present invention will be shown.
実施例１゜
バチルス・ズブチリスＮｏ、０６０．ＦＥＲＭ Ｐ−８
５４２を次の培地に接種して、２７℃で４日間振どう培
養した。Example 1 Bacillus subtilis No. 060. FERM P-8
542 was inoculated into the following medium and cultured with shaking at 27°C for 4 days.
ポリペプトン ３０ｇ
酵母エキス ５ｇ
ＮａＣＱ ５ ｇ
脱イオン水 ＩＱ
（ｐ）Ｉ ７．０）
得られた麹養液を濾過して、培養濾液とし、これを６Ｎ
Ｈ（、Ｑでｐ）Ｉ ＝ ３．０として、しばらく放置
すると沈澱物が得られるので、得られた沈澱物を蒸留水
に溶解し、これにＮａ）Ｉｃｏ、を加え、 ｐＨ＝７．
０とし、更にエタノールを８０％まで加え、濾過する。Polypeptone 30g Yeast extract 5g NaCQ 5g Deionized water IQ (p)I 7.0) The obtained malt solution was filtered to obtain a culture filtrate, and this was 6N
When H(, p in Q)I = 3.0, a precipitate will be obtained if it is left to stand for a while, so the obtained precipitate is dissolved in distilled water, Na)Ico is added thereto, and pH = 7.
0, then add ethanol to 80% and filter.
得られた上清を真空乾燥し、乾固物を得る。この乾固物
を蒸留水に溶解し、６Ｎ ＩＩｃＱでＰＨ＝３．０とし
、しばらく放置し、沈澱物を得る。この沈澱物を蒸留水
に溶解し、ＮａｔｌＣＯｉでｐｌ＋ ＝ ７．０とし、
この溶液を蒸留水に対して透析する。The obtained supernatant is vacuum dried to obtain a dried product. This dried product is dissolved in distilled water, adjusted to pH=3.0 with 6N IIcQ, and left to stand for a while to obtain a precipitate. This precipitate was dissolved in distilled water and adjusted to pl+ = 7.0 with NatlCOi,
This solution is dialyzed against distilled water.
得られた透析液を凍結乾燥し、Ｐ４物質含有の粗精製物
を黄色粉末で得る。The obtained dialysate is freeze-dried to obtain a crude product containing the P4 substance as a yellow powder.
得られた黄色粉末を高速液体クロマトにかけ。The resulting yellow powder was subjected to high performance liquid chromatography.
第２図に矢印で示すＰ４物質該当部分をとり、これを凍
結乾燥することにより、はぼ純品のＰ４物質を白色無定
形粉末として得た。A portion corresponding to the P4 substance indicated by the arrow in FIG. 2 was taken and freeze-dried to obtain pure P4 substance as a white amorphous powder.
第１図はＰ４物質の紫外線吸収スペクトルを示す図で、
第２図はＰ４物貿含有粗精製物の葛速液体クロマトによ
る人２１０による吸光度曲線を示す図である。矢印はＰ
４物質を示す。
代理人 弁理士 戸 １）親 男
第 ２ 図
開聞 （分）Figure 1 shows the ultraviolet absorption spectrum of P4 substance.
FIG. 2 is a diagram showing an absorbance curve of a crude product containing P4 obtained by human 210 using Kuzuhaku liquid chromatography. The arrow is P
Shows 4 substances. Agent Patent Attorney 1) Parent Male 2nd Dialogue (Minutes)
Claims (3)
不溶で、希エタノール水、希メタノール水、ＣＨＣｌ＿
３に可溶である。 ３、物質の色、性状：白色無定形粉末 ４、呈色反応：ニンヒドリンテスト ５、本物質のアミノ酸組成：本物質の加水分解物のアミ
ノ酸分析によって次の結果を得た。 ６、本物質は真菌に対して強い抗菌性を示す。 サッカロミセス・セレビシエＩＡＭ４１２５の生育を１
６０μｇ／ｌで阻止する。(1) P4 substance having the following physical and chemical properties. 1. The ultraviolet absorption spectrum is as shown in FIG. 2. Solubility: Insoluble in ethanol, methanol, acetone, water, dilute ethanol water, dilute methanol water, CHCl_
It is soluble in 3. 3. Color and properties of the substance: White amorphous powder 4. Color reaction: Ninhydrin test 5. Amino acid composition of this substance: The following results were obtained by amino acid analysis of the hydrolyzate of this substance. 6. This substance exhibits strong antibacterial properties against fungi. Growth of Saccharomyces cerevisiae IAM4125 1
Inhibit at 60 μg/l.
４物質を採取することを特徴とするＰ４物質の製造法。(2) Cultivate P4 substance-producing bacteria belonging to the genus Bacillus, and
A method for producing a P4 substance, which comprises collecting four substances.
ズブチリスである特許請求の範囲第２項記載の製造法。(3) P4 substance-producing bacteria belonging to the genus Bacillus are Bacillus.
The manufacturing method according to claim 2, which is S. subtilis.
Priority Applications (1)
| Application Number | Priority Date | Filing Date | Title |
|---|---|---|---|
| JP60275220A JPS62135497A (en) | 1985-12-09 | 1985-12-09 | P4 substance and preparation thereof |
Applications Claiming Priority (1)
| Application Number | Priority Date | Filing Date | Title |
|---|---|---|---|
| JP60275220A JPS62135497A (en) | 1985-12-09 | 1985-12-09 | P4 substance and preparation thereof |
Publications (1)
| Publication Number | Publication Date |
|---|---|
| JPS62135497A true JPS62135497A (en) | 1987-06-18 |
Family
ID=17552374
Family Applications (1)
| Application Number | Title | Priority Date | Filing Date |
|---|---|---|---|
| JP60275220A Pending JPS62135497A (en) | 1985-12-09 | 1985-12-09 | P4 substance and preparation thereof |
Country Status (1)
| Country | Link |
|---|---|
| JP (1) | JPS62135497A (en) |
Cited By (1)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| JPS62135496A (en) * | 1985-12-09 | 1987-06-18 | Kibun Kk | P3 substance and preparation thereof |
Citations (1)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| JPS62135496A (en) * | 1985-12-09 | 1987-06-18 | Kibun Kk | P3 substance and preparation thereof |
-
1985
- 1985-12-09 JP JP60275220A patent/JPS62135497A/en active Pending
Patent Citations (1)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| JPS62135496A (en) * | 1985-12-09 | 1987-06-18 | Kibun Kk | P3 substance and preparation thereof |
Cited By (1)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| JPS62135496A (en) * | 1985-12-09 | 1987-06-18 | Kibun Kk | P3 substance and preparation thereof |
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