pH和色谱柱对日本鳗鲡肝脏抗菌肽分离纯化效果的影响及抗菌活性检测
Effects of pH and chromatographic column on isolation, purification of antibacterial peptide from Japanese eel liver, and determination of antibacterial activity
查看参考文献17篇
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文摘
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为了能够快捷地分离纯化出单一的抗菌肽,实验采用不同pH值(离子交换流动相pH3.0、4.0、5.0;反相液相层析流动相pH 2.0、4.5、7.0)缓冲液,不同色谱柱(离子交换和凝胶过滤层析柱分别与反相液相层析柱)联用对日本鳗鲡肝脏抗菌肽分离纯化效果进行了比较,并分析了琼脂板扩散法和微孔液体培养法检测抗菌活性的优缺点。结果显示:pH 4.0缓冲液为最佳离子交换流动相,蛋白提取率为16.43%,得到2个洗脱峰;反相液相层析3种pH缓冲液分离效果均不理想,凝胶过滤层析柱与反相液相层析柱联用能明显改善日本鳗鲡肝脏抗菌肽分离纯化效果,洗脱峰数量较多,峰形单一且尖锐狭窄,基线平而低。琼脂板扩散法检测抗菌活性操作简单,实验结果直观,但所需蛋白量较多,可用于蛋白粗提物抗菌活性的检测;微孔液体培养法检测抗菌活性灵敏,所需蛋白较少,可用于后期色谱分离纯化蛋白抗菌活性的检测。 |
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其他语种文摘
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In order to quickly isolate and purify antimicrobial peptide,different flow phase pH values(cationchange pH 3. 0,4. 0,5. 0,and reverse-phase liquid chromatography pH 2. 0,4. 5,7. 0), and different chromatographic columns (cation-change and gel chromatography coupled with reverse-phase liquid chromatography respectively) were used to isolate and purify antibacterial peptide from Anguilla japonica liver,and the results were analysed and compared. The merits and drawbacks of agar plate diffusion and microporous liquid culture methods on determination of antibacterial activity were analysed. The results showed that pH 4. 0 buffer solution was the best flow phase in three-pH-value buffer solutions,which had 16. 43% extraction ratio and isolated two elution peaks; We did not obtain perfect result from reverse-phase liquid chromatography (RPC) in three-pH-value buffer solutions; Sephadex G-25 gel coupled with RPC significantly improved the isolation effect,elution peak amounts were greater,peaks were sharper and narrower,baseline was flatter and low than those of cation exchange coupled with RPC. Agar plate diffuse method to determine antibacterial activity was simple to operate,and experimental result was intuitional. It required much protein, so it should be used to determine antibacterial activity during protein crude extraction period; Microporous liquid culture method to determine antibacterial activity was sensitive. It required little protein, so it should be used to determine antibacterial activity during isolation and purification of protein in chromatography period. |
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来源
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水产学报
,2013,37(4):614-621 【核心库】
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关键词
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日本鳗鲡
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抗菌肽
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pH
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色谱柱
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抗菌活性
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地址
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1.
集美大学水产学院, 鳗鲡现代产业技术教育部工程研究中心, 福建, 厦门, 361021
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鑫双宝生物科技(厦门)有限公司, 福建, 厦门, 361021
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语种
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中文 |
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文献类型
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研究性论文 |
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ISSN
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1000-0615 |
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学科
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水产、渔业 |
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基金
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国家公益性行业（农业）科研专项
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福建省教育厅科技项目
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厦门市工程技术研究中心创新项目
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厦门市科技局重点项目
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文献收藏号
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CSCD:4805057
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