洋葱伯克霍尔德菌CF-66环二肽的分离纯化及抑菌活性研究  

Purification and Antifungal Characterization of a Diketopiperazine from Burkholderia cepacia CF-66

作　　者：王艾晶[1,2] 权春善[3] 王建华[1,2] 王军华[3] 范圣第[3] 

机构地区：[1]中国科学院大连化学物理研究所,大连116023 [2]中国科学院研究生院,北京100049 [3]大连民族学院生命科学学院,大连116600

出　　处：《中国生物工程杂志》2009年第12期43-48,共6页China Biotechnology

基　　金：辽宁省教育厅资助项目(223038)

摘　　要：植物真菌病害给农业生产带来了巨大损失,因此对高效、低毒、低残留的生物农药的开发迫在眉睫。洋葱伯克霍尔德菌CF-66(Burkholderia cepacia CF-66)对真菌类病原菌具有强烈的抑制作用。其发酵液经减压浓缩和乙酸乙酯萃取得到粗提液,粗提液经反复硅胶柱层析和反相高效液相色谱(RP-HPLC)多步柱层析,首次分离纯化得到一种环二肽——cyclo(Phe-Pro)(cFP)。利用气质联用(GC-MS)系统和HPLC进行定性和定量,结果表明分离纯化物质呈单峰,纯度较高且经标准曲线算出其浓度约为15 mg/ml。MIC值的测定结果表明该物质对立枯丝核菌、黄瓜菌核、玉米弯孢病菌等植物病原菌及冻土毛霉、黄曲霉、米根霉等食品腐败菌均具有较强的抑制作用。经显微镜观察发现,该物质可使丝状真菌菌丝生长异常,菌丝由光滑细长变得粗糙、弯曲、短粗且顶端膨大呈泡状。Fungal diseases cause a considerable damage to many economic crops,while chemical control of diseases has a huge threat to human health and the environment. Therefore, it is becoming extremely urgent to develop biological pesticide with high activity,low toxicity and low residues. Burkholderia cepacia CF-66, isolated from the compost, shows strong antifungal activity due to various secondary metabolites, such as pyrrolnitrin, phenazine, diketopiperazines, siderophores and so on. One of diketoperazines, namely, cyclo （ Phe-Pro ） （cFP） was purified through the following steps. Firstly the crude extract was obtained from the fermentation broth of B. cepacia CF-66 by centrifugation,reduced pressure distillation and ethyl acetate extraction. Then cFP was isolated and purified by silica gel column chromatography and RP-HPLC. Three peaks were exculsively detected. The third one（cFP） was collected and analyzed by HPLC and GC-MS, showing single peak with a concentration of 15mg/ml. Finally, the antifungal activity was carried out by detecting minimal inhibitory concerntration （ MIC ）, which demonstrated that cFP significantly inhibited the growth of Sderotinia sclerotiorum, Rhhoctonia solani kvhn 0-28, Curvularia lunata and so forth. The result of microscopic observation showed that cFP could cause twisted hypha,fraetured hypha and swollen tips during hyphal development. In conclusion, cFP has great potential for developing biological pesticide.

关 键 词：洋葱伯克霍尔德菌 环二肽 分离纯化 抑菌性 

分 类 号：Q819[生物学—生物工程]