CN113186111A - 一株深海放线菌红串红球菌及其抑制黄曲霉毒素的应用 - Google Patents
一株深海放线菌红串红球菌及其抑制黄曲霉毒素的应用 Download PDFInfo
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Abstract
本发明提供一株可抑制黄曲霉毒素合成的深海放线菌菌株，为红串红球菌BC14‑M2AF‑1菌株(Rhodococcus erythropolis BC14‑M2AF‑1)，分离自西太平洋6105米水深的沉积物，已保藏于广东省微生物菌种保藏中心，保藏编号为GDMCC No.61239。将BC14‑M2AF‑1菌株在淡水M2培养基中发酵，获得的发酵无细胞上清液可从黄曲霉毒素合成途径的源头100%抑制黄曲霉毒素；发酵无细胞上清液在121度、103kPa下高温高压处理30分钟，仍能100%抑制黄曲霉毒素的合成。本发明的红串红球菌BC14‑M2AF‑1菌株及其发酵产物可用于田间作物栽培、粮食储藏、食品与饮料加工、饲料生产等的黄曲霉毒素污染的有效防控。
Description
技术领域：
本发明涉及深海生物技术与黄曲霉毒素生物防控技术领域，具体的说是一株深海放线菌红串红球菌及其抑制黄曲霉毒素的作用。
背景技术：
黄曲霉毒素是被国际癌症机构列为I类致癌物质的真菌毒素，主要由黄曲霉、寄生曲霉等曲霉属真菌产生，不仅可广泛污染田间种植的粮食，而且在粮食储藏期间、及由其加工而成的食品和饲料，同样存在黄曲霉毒素的污染，人类和动物食用被黄曲霉毒素污染的食物和饲料后，可严重危害人类和动物的健康，因此，有效抑制黄曲霉毒素在粮食、饲料等的污染是全球亟待解决的重大食品和饲料安全问题，具有重要的现实意义、显著的经济和社会效益。
在陆地资源日益匮乏的今天，世界各国将目标转向海洋，尤其是大洋深海等极端环境的微生物资源及其开发利用成为世界瞩目与竞争的焦点，开发深海微生物资源在黄曲霉毒素污染防控方面的研究与应用，在世界上尚不多见。
发明内容：
本发明所要解决的技术问题是克服上述现有技术的不足，所采用的技术方案是：一株深海放线菌红串红球菌及其抑制黄曲霉毒素的作用，其特征包括如下步骤：
（1）本发明所说的深海放线菌是红串红球菌BC14-M2AF-1菌株(RhodococcuserythropolisBC14-M2AF-1) ，并已于2020年10月26日保藏于广东省微生物菌种保藏中心，保藏编号为GDMCC No.61239；保藏地址广东省广州市先烈中路100号大院59号楼5楼。
（2）本发明所说的红串红球菌BC14-M2AF-1菌株分离自西太平洋6105米水深的沉积物，通过以下方法分离获得：将4度保存的沉积物10倍系列稀释后，涂布于淡水M2培养基，28度下恒温培养，及时挑取单菌落，并经进一步划线纯化，获得纯菌种BC14-M2AF-1菌株；
（3）本发明所说的BC14-M2AF-1菌株经16S rRNA基因分子鉴定为红串红球菌(Rhodococcus erythropolis) 的一个新菌株；
（4）将本发明的红串红球菌BC14-M2AF-1菌株接种于淡水配制的M2培养基中，在18-32℃温度下，振荡培养4-10天，即可得到含有高效抑制黄曲霉毒素合成的活性物质发酵液，该发酵液的无细胞上清液可100%抑制黄曲霉毒素的合成。
（5）本发明的红串红球菌BC14-M2AF-1菌株的发酵液中的活性物质具有高温高压稳定性，在121℃、103kPa的高温高压下处理30分钟后活性不丧失；
（6）本发明的红串红球菌BC14-M2AF-1菌株及其发酵液可应用于田间作物栽培、粮食储藏、食品与饮料加工、饲料生产等的黄曲霉毒素污染的有效防控。
具体实施方式
下面结合实施例对本发明做进一步说明，但并不因此将本发明限制在所述的实施例范围之中。
实施例1：红串红球菌BC14-M2AF-1菌株的发酵
将红串红球菌BC14-M2AF-1菌株的液体种子，按1-8%接种量接种到含有乙酸钠0.5%，蛋白胨 0.05%，酵母浸粉0.05%，牛肉膏0.05%，葡萄糖 0.05%，蔗糖 0.05%，可溶性淀粉 0.05%，柠檬酸三钠 0.005%，苹果酸 0.005%，酒石酸钾钠 0.005%，硝酸铵 0.1%，氯化铵0.02%、淡水100ml的液体培养基中，在18-32℃温度、100-240转/分钟下培养4-10天后，对发酵液进行离心，获得的无细胞上清液，即是含有高效抑制黄曲霉毒素合成的活性物质的上清液，可直接用于抑制真菌黄曲霉毒素的合成，或者加工成粉剂用于抑制真菌黄曲霉毒素的合成。
实施例2：红串红球菌BC14-M2AF-1菌株发酵无细胞上清液对黄曲霉毒素合成的抑制作用
将实施例1获得的无细胞发酵上清液中，按照2%葡萄糖和0.5%酵母膏的配比，加入到发酵上清液中以补充营养，作为培养黄曲霉毒素产生菌-寄生曲霉的培养液，对照培养液为实施例1中的液体培养基中加入2%葡萄糖和0.5%酵母膏，向上述培养基中分别接种1%的寄生曲霉的孢子悬液后，在28度培养6天，离心收集菌丝体，用甲醇-氢氧化钠溶液提取菌丝体中的黄曲霉毒素生物合成途径的中间产物，在560nm处测量光密度值（OD560），计算抑毒率，抑毒率(%)=(对照培养液的OD560-无细胞发酵上清液的OD560)/对照培养液的OD560 x100，经测定和计算，红串红球菌BC14-M2AF-1菌株发酵无细胞上清液对黄曲霉毒素合成的抑制率达100%。
实施例3：红串红球菌BC14-M2AF-1菌株发酵液中活性物质的高温高压稳定性
将实施例1获得的无细胞发酵上清液在121oC、103kPa下高温高压处理30分钟后自然冷却至室温，将上述处理的发酵液、未经过温度处理的发酵液、和对照培养液一起分别接种1%的寄生曲霉的孢子悬液后，在28度培养6天，离心收集菌丝体，用甲醇-氢氧化钠溶液提取菌丝体中的黄曲霉毒素生物合成途径的中间产物，在560nm处测量光密度值（OD560），计算抑毒率，抑毒率(%)=(对照培养液的OD560-无细胞发酵上清液的OD560)/对照培养液的OD560 x100，经测定和计算，即使在121oC、103kPa下高温高压处理30分钟的发酵液也能与未经过温度处理的发酵液一样，对黄曲霉毒素合成的抑制率达100%，表明发酵液中的抑毒活性物质具有高温高压稳定性。
Claims (1)
1.一株分离自西太平洋6105米水深的沉积物的深海放线菌菌株，为红串红球菌BC14-M2AF-1菌株 (Rhodococcus erythropolis BC14-M2AF-1)，已于2020年10月26日保藏于广东省微生物菌种保藏中心，保藏编号为GDMCC No. 61239。
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Citations (4)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| WO2002099142A2 (de) * | 2001-06-01 | 2002-12-12 | Wilhelm Holzapfel | Actinomyceten zum abbau von aflatoxin b1, ochratoxin a und/oder zearalenon |
| CN104738363A (zh) * | 2013-12-27 | 2015-07-01 | 中粮营养健康研究院有限公司 | 红城红球菌在降解饲料或其原料中的黄曲霉毒素b1中的用途 |
| CN104745493A (zh) * | 2013-12-27 | 2015-07-01 | 中粮营养健康研究院有限公司 | 降解黄曲霉毒素b1的红城红球菌菌株的分离、培养和使用方法 |
| CN105039199A (zh) * | 2015-06-10 | 2015-11-11 | 哈尔滨工业大学（威海） | 一株深海环状芽孢杆菌及其抑制黄曲霉毒素的应用 |
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2020
- 2020-12-15 CN CN202011473063.0A patent/CN113186111A/zh active Pending
Patent Citations (4)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| WO2002099142A2 (de) * | 2001-06-01 | 2002-12-12 | Wilhelm Holzapfel | Actinomyceten zum abbau von aflatoxin b1, ochratoxin a und/oder zearalenon |
| CN104738363A (zh) * | 2013-12-27 | 2015-07-01 | 中粮营养健康研究院有限公司 | 红城红球菌在降解饲料或其原料中的黄曲霉毒素b1中的用途 |
| CN104745493A (zh) * | 2013-12-27 | 2015-07-01 | 中粮营养健康研究院有限公司 | 降解黄曲霉毒素b1的红城红球菌菌株的分离、培养和使用方法 |
| CN105039199A (zh) * | 2015-06-10 | 2015-11-11 | 哈尔滨工业大学（威海） | 一株深海环状芽孢杆菌及其抑制黄曲霉毒素的应用 |
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