家蝇天然抗菌肽的分离纯化与活性分析

Purification and Activity Analysis of Antibacterial Peptide from House Fly

目的:纯化得到有抗菌活性的家蝇天然抗菌肽。方法:以革兰氏阴性菌(大肠杆菌)和革兰氏阳性菌(金黄色葡萄球菌)的混合菌液,在未灭活的情况下对家蝇幼虫实施带菌针刺以诱导抗菌肽的大量表达。采用固相萃取(SPE),得到三个对大肠杆菌和金黄色葡萄球菌均有强烈抑制作用的组分Sp1、Sp3和Sp8。用反相高效液相色谱(RP-HPLC)对活性最强的Sp1进行了纯化,液体生长抑制法证明其中的组分Ⅲ具有最强活性。毛细管电泳(CE)显示该组分由两种物质组成。结果和结论:家蝇血淋巴中存在大量有抗菌活性的物质。该研究利用SPE和RP-HPLC与CE联用的方法,得到了有较强抗菌活性和较宽抗菌谱的粗提物,并获得一个推测由两种阳离子型抗菌肽组成的成分,为后期研究打下了基础。

Objective： To get the natural purified antibacterial peptides with the activity of antibacterium from house fly（Musca domestica）.Methods： To inject the instars with living bacteria mixture of Gram negative（Escherichia coli） and Gram positive bacterium（Staphylococcus aureus） to induce the abundant expression of antibacterial peptides in the body.Hemolymph from house fly was pretreated with solid phase extraction（SPE）,which produced three components such as Sp1,Sp3 and Sp8 that showed great resistance to E.coli and S. aureus. Spl with the greatest activity was isolated with RP- HPLC. The further analysis showed that the peak Ⅲ of RP- HPLC has a top activity through liquid growth inhibition method. And the results based on capillary electrophoresis proved that the peak Ⅲ has two components. Results and Conclusion： There are a great deal of material with the activity of antibacterium. Here, we have obtained the active primary extraction of wide antibacterial spectrum from the hemolymph of house fly, and a component possibly composed of two cationic antibacterial peptides by SPE, RP - HPLC and CE, which set a good foundation for the further research.