日本鳗鲡脾脏抗菌肽抗菌活性检测方法的比较

Comparative Study on Test Methods for Antibacterial Activities of Antibacterial Peptides from Spleen of Japanese Eel( Anguilla japonica)

本试验旨在建立一种在日本鳗鲡脾脏抗菌肽分离纯化过程中简便、灵敏的检测其抗菌活性的方法。试验以日本鳗鲡脾脏分子质量小于10 ku的蛋白质为抗菌肽样品,选择琼脂板孔穴扩散法和微孔液体培养法2种传统方法及微量液体培养法为检测方法,比较这3种方法在抗菌肽对3种鳗鲡常见致病菌株(迟钝爱德华菌、气单胞菌、嗜水气单胞菌)抗菌活性检测中的灵敏度和优缺点。结果表明:与琼脂板孔穴扩散法及微孔液体培养法相比,微量液体培养法简便、易于观察结果,具有最高的灵敏度且样品用量最少,适合在鳗鲡脾脏抗菌肽分离纯化过程中跟踪检测抗菌活性,尤其适合分离纯化后期获得微量单一样品抗菌活性的检测。由此得出,微量液体培养法适用于检测日本鳗鲡脾脏抗菌肽的抗菌活性。

The study was to establish a simple and sensitive test method for antibacterial activity analysis in the separation and purification of antibacterial peptides from the spleen of Japanese eel（ Anguilla japonica）. Experiment by using spleen protein of molecular mass 10 ku from Japanese eel as antibacterial peptide samples,to compare the sensitivity,advantages and disadvantages of three test methods. In these three test methods,two conventional methods were agar plate fovea diffusion method and microporous liquid culture method and another was trace liquid culture method,for testing antibacterial activities of antibacterial peptides against three common pathogenic bacterial strains of eel,including Edwardsiella tarda,Aeromonas sp. and Aeromonas hydrophila. The results showed that compared with the agar plate fovea diffusion method and microporous liquid culture method,the trace liquid culture method had the advantages of simple operation and easy to observe result. Furthermore,this method with the best sensitivity and using the least sample quantity could be suitable for testing antibacterial activities in the separation and purification of antibacterial peptides from the spleen of eel,especially for identifying antibacterial activity of the a fewand single fraction obtained in late stage of separation and purification. This study suggests that the trace liquid culture method is suitable for testing antibacterial activities of antibacterial peptides from the spleen of Japanese eel.