伯克霍尔德菌HQB-1抑制香蕉枯萎病菌的活性化合物分离鉴定

Isolation and identification of bioactive compounds of Burkholderia HQB-1 strain inhibiting banana wilt

【背景】伯克霍尔德菌HQB-1对香蕉枯萎病菌(Fusariumoxysporumf.sp.cubenseTropical Race 4,Foc TR4)具有良好的防治效果。【目的】从菌株HQB-1发酵液中分离活性化合物并通过超高效液相联用质谱进行检测,获得该菌株具有良好生防作用的单体化合物。【方法】以HQB-1为目标菌株,大批量发酵并进行发酵液分离、提纯,通过超高效液相联用质谱法与核磁共振波谱法鉴定活性化合物。【结果】HQB-1菌株发酵液中的蛋白对Foc TR4无抑制作用;HQB-1菌株产生儿茶酚型铁载体,而不产生异羟肟酸型铁载体;对HQB-1菌株发酵液离心、浓缩、干燥,获得乙酸乙酯浸膏(粗提物),经过大孔树脂柱的充分吸附,在30%、60%及无水甲醇的洗脱下获得组分1-3,对Foc TR4的抑菌率分别为10.06%、27.82%和51.40%;选择抑菌率最大的组分3过硅胶柱层析,获得黄绿色晶体。该化合物在365 nm波长下具有最大吸收峰,将核磁共振图谱与SciFinder和SDBS信息数据库进行图谱比对,将该抑菌活性化合物鉴定为吩嗪-1-羧酸(Phenazine-1-Carboxylic Acid,PCA)。PCA对Foc TR4的最小抑菌浓度最低,仅为1.563μg/mL,说明PCA对Foc TR4的抑制效果较强。【结论】从HQB-1菌株中分离得到活性化合物PCA,PCA的发现为香蕉枯萎病的生物防治奠定了良好的理论基础。

[Background] Burkholderia HQB-1 has good control effect on banana fusarium wilt(Fusarium oxysporum f. sp. cubense tropical race 4(Foc TR4)). [Objective] The active compound was separated from the fermentation broth of HQB-1 strain, and the active compound was detected by HPLC-MS to obtain the monomer compound with good biocontrol effect. [Methods] HQB-1 strain was fermented in large quantities, and then the fermentation broth was separated and purified. The active compounds were identified by HPLC-MS and NMR. [Results] The protein from fermentation broth of HQB-1 strain had no inhibitory effect on Foc TR4. The catechol type iron carrier rather than hydroxamic acid type iron carrier was produced from HQB-1 strain. HQB-1 strain fermentation broth was centrifuged, concentrated and dried to obtain ethyl acetate extract(crude extract), which was then fully adsorbed by macroporous resin column. The components 1-3 were obtained after eluting by 30%, 60% and anhydrous methanol, and the corresponding inhibitory rate to Foc TR4 was 10.06%, 27.82% and 51.40% respectively. The component with the highest inhibitory rate was selected, and the yellow green crystal was obtained through silica gel column chromatography. The compound had a maximum absorption peak at 365 nm wavelength. The antibacterial activity compound was identified as phenazine-1-carboxylic acid(PCA), after the comparative analysis of NMR spectrum with SciFinder and SDBS information database. The minimum inhibitory concentration(MIC) of PCA to Foc TR4 was the lowest, only 1.563 μg/mL, indicating that PCA had a strong inhibitory effect on Foc TR4. [Conclusion] The active compound PCA isolated from HQB-1 strain had the biocontrol effect on banana fusarium wilt.