CN106146579B - 一种提纯阿维拉霉素的方法 - Google Patents

一种提纯阿维拉霉素的方法 Download PDF

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CN106146579B
CN106146579B CN201610663519.7A CN201610663519A CN106146579B CN 106146579 B CN106146579 B CN 106146579B CN 201610663519 A CN201610663519 A CN 201610663519A CN 106146579 B CN106146579 B CN 106146579B
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avilamycin
crude product
alkaline
methanol
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CN106146579A (zh
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周超
陈文成
陶锐
孙春领
尤敏
张胜龙
师丽君
韩振国
刘远声
李玄涛
郭彦超
周志霞
陈志�
孙玉佶
高淑玲
王慧慧
张嫄婧
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PUYANG HOTWAY PHARMACEUTICALS CO Ltd
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    • C07H—SUGARS; DERIVATIVES THEREOF; NUCLEOSIDES; NUCLEOTIDES; NUCLEIC ACIDS
    • C07H17/00—Compounds containing heterocyclic radicals directly attached to hetero atoms of saccharide radicals
    • C07H17/04—Heterocyclic radicals containing only oxygen as ring hetero atoms
    • C—CHEMISTRY; METALLURGY
    • C07—ORGANIC CHEMISTRY
    • C07H—SUGARS; DERIVATIVES THEREOF; NUCLEOSIDES; NUCLEOTIDES; NUCLEIC ACIDS
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    • C07H1/06—Separation; Purification

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Abstract

本发明涉及一种抗生素阿维拉霉素提纯的方法。该方法是以甲醇溶剂浸提含阿维拉霉素的菌丝体，然后通过结晶、甲醇钠反应、碱性氧化铝柱层析、重结晶等步骤获得阿维拉霉素A和B纯品。本发明最终得到的纯品分别是阿维拉霉素A和阿维拉霉素B的纯品，纯化步骤简便，易工业化，平均得率在42.5％以上，纯度在98.5％以上。

Description

一种提纯阿维拉霉素的方法
技术领域
本发明涉及一种抗生素阿维拉霉素提纯的方法，属于生物医药、饲料添加剂领域。
背景技术
阿维拉霉素(Avilamycin)结构特殊，一般由绿色产色链霉菌(Streptomycesviridoehrongenes)发酵而成，属于正糖霉素族的寡糖类抗生素，它主要抑制革兰氏阳性菌，是一种新型的抗菌促生长剂和代谢调节剂。在畜禽业中，因其安全性高、不存在交叉耐药性及良好稳定的促生长效果，越来越受到人们的重视。阿维拉霉素是一种混合型低聚糖类抗生素，目前发现有A、B、C、D、F、G、H等多种结构，其中最具活性的是阿维拉霉素A组分和B组分，其他成分活性很小，并且含量很低。
现有阿维拉霉素的生产技术被美国礼来一家公司垄断，国内在高单位的阿维拉霉素菌种选育及工艺研究上仍处于研发阶段，提纯研究也很少，技术很不成熟，市场上阿维拉霉素的纯品几乎没有。CN103012519A通过结晶、硅胶柱层析、ODS柱洗脱分离、重结晶、等步骤获得阿维拉霉素A和B纯品,提纯步骤复杂且需要两次柱层析，不利于工业化生产。
发明内容
本发明要解决的问题是：克服现有技术中阿维拉霉素提纯技术的不足之处，提出一种可行的阿维拉霉素提纯方法。
为解决该技术问题，本发明的技术方案是：
提供一种阿维拉霉素提纯的方法，包括以下步骤：
(1)将含有阿维拉霉素的菌丝体加入甲醇中，室温下超声，离心得上清液，重复上述步骤三次，合并上清夜即得浸提液，按浸提液∶水的体积比为1∶1-1∶2进行补水结晶，离心，烘干得阿维拉霉素粗提物；
(2)阿维拉霉素粗提物用甲醇溶解，冰浴下按照甲醇钠与阿维拉霉素粗提物质量比为1∶6-1∶12分批添加甲醇钠，搅拌30-45min，加入阿维拉霉素粗提物重量3-5倍的碱性氧化铝，减压浓缩得拌样粗品；
(3)将拌样粗品干法上样碱性氧化铝柱，用氯仿与甲醇体积比为96：4洗脱，分别收集阿维拉霉素A和阿维拉霉素B的洗脱液，减压浓缩得到阿维拉霉素A碱性粗品和阿维拉霉素B碱性粗品；
(4)将阿维拉霉素A碱性粗品和阿维拉霉素B碱性粗品分别用甲醇溶解，用1mol/LHCl溶液调节PH为6-8，按照阿维拉霉素A碱性粗品和阿维拉霉素B碱性粗品溶液∶水的体积比为1∶1-1∶2分别进行加水重结晶，抽滤后结晶用纯水漂洗，烘干后即获得阿维拉霉素A和阿维拉霉素B纯品。
本发明所得阿维拉霉素A和阿维拉霉素B的纯品的平均得率为42.5％以上，并且阿维拉霉素A和阿维拉霉素B的纯度在98.5％以上。
技术效果本发明与现有技术相比，具有如下特点：
本发明使用甲醇钠和阿维拉霉素粗提物反应，增大阿维拉霉素A和阿维拉霉素B的极性的同时，阿维拉霉素A和阿维拉霉素B保持稳定性，有利于进一步提纯；
本发明采用一次柱层析，两次结晶的提纯方法，大大简化了提纯步骤，有利于工业化生产。
具体实施方式
实施例1：
将5g含量为20％阿维拉霉素的菌丝加入100mL甲醇中，室温下超声90min，离心得上清液，重复上述步骤三次，合并上清夜即得浸提液，按浸提液∶水的体积比为1∶1进行补水结晶，离心，烘干得阿维拉霉素粗提物。
阿维拉霉素粗提物用甲醇溶解，冰浴下按照甲醇钠与阿维拉霉素粗提物质量比为1∶12分批添加甲醇钠，搅拌30min，加入阿维拉霉素粗提物重量3倍的碱性氧化铝，减压浓缩得拌样粗品；
将拌样粗品干法上样碱性氧化铝柱，用氯仿与甲醇体积比为96：4洗脱，分别收集阿维拉霉素A和阿维拉霉素B的洗脱液，减压浓缩得到阿维拉霉素A和阿维拉霉素B碱性粗品；
将阿维拉霉素A和阿维拉霉素B碱性粗品分别用甲醇溶解，用1mol/L HCl溶液调节PH为7，按照阿维拉霉素A和阿维拉霉素B碱性粗品溶液∶水的体积比为1∶1分别进行加水重结晶，抽滤后结晶用纯水漂洗，烘干后即获得阿维拉霉素A纯品0.33g和阿维拉霉素B纯品0.10g。

Claims (2)

1.一种阿维拉霉素提纯的方法，其特征在于，包括以下步骤：
(1)将含有阿维拉霉素的菌丝体加入甲醇中，室温下超声，离心得上清液，重复上述步骤三次，合并上清夜即得浸提液，按浸提液∶水的体积比为1∶1-1∶2进行补水结晶，离心，烘干得阿维拉霉素粗提物；
(2)阿维拉霉素粗提物用甲醇溶解，冰浴下按照甲醇钠与阿维拉霉素粗提物质量比为1∶6-1∶12分批添加甲醇钠，搅拌30-45min，加入阿维拉霉素粗提物重量3-5倍的碱性氧化铝，减压浓缩得拌样粗品；
(3)将拌样粗品干法上样碱性氧化铝柱，用氯仿与甲醇体积比为96：4洗脱，分别收集阿维拉霉素A和阿维拉霉素B的洗脱液，减压浓缩得到阿维拉霉素A碱性粗品和阿维拉霉素B碱性粗品；
(4)将阿维拉霉素A碱性粗品和阿维拉霉素B碱性粗品分别用甲醇溶解，用1mol/L HCl溶液调节pH为6-8，按照阿维拉霉素A碱性粗品和阿维拉霉素B碱性粗品溶液∶水的体积比为1∶1-1∶2分别进行加水重结晶，抽滤后结晶用纯水漂洗，烘干后即获得阿维拉霉素A和阿维拉霉素B纯品。
2.根据权利要求1所述的一种阿维拉霉素提纯的方法，其特征在于，步骤(4)中pH优选调节为7。
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Citations (3)

* Cited by examiner, † Cited by third party
Publication number Priority date Publication date Assignee Title
EP0118968A1 (en) * 1983-01-17 1984-09-19 Eli Lilly And Company Use of avilamycin to control swine dysentery
CN101817745A (zh) * 2009-12-29 2010-09-01 华中农业大学 一种3,5-二氯-4-羟基-2-甲氧基-6-甲基-苯甲酸的化学合成方法
CN103012519A (zh) * 2012-12-13 2013-04-03 浙江凯胜科技有限公司 一种兽用抗生素阿维拉霉素分离纯化的方法

Patent Citations (3)

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EP0118968A1 (en) * 1983-01-17 1984-09-19 Eli Lilly And Company Use of avilamycin to control swine dysentery
CN101817745A (zh) * 2009-12-29 2010-09-01 华中农业大学 一种3,5-二氯-4-羟基-2-甲氧基-6-甲基-苯甲酸的化学合成方法
CN103012519A (zh) * 2012-12-13 2013-04-03 浙江凯胜科技有限公司 一种兽用抗生素阿维拉霉素分离纯化的方法

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