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【目的】应用高速逆流色谱法(High-speed counter-current chromatography,HSCCC)实现对小孢拟盘多毛孢Pestalotiopsis microspora PM-1菌株代谢产物中除草活性物质的首次分离。【方法】以正己烷:乙酸乙酯:甲醇:水(4:5:4:5,V/V/V/V)为最佳的两相溶剂体系,上相(水相)为固定相,下相(有机相)为流动相,正相洗脱。【结果】在流速2 mL/min、转速900 r/min、检测波长254 nm的条件下进行分离,得到4个馏分,其中馏分Ⅱ对马唐的活性较强。馏分Ⅱ经高效液相色谱法(High performance liquid chromatography,HPLC)以乙腈:水=75:25(V/V)为流动相经过C18柱进一步分离检测,所得馏分A对马唐种子萌发有较强的抑制作用。其保留时间为7.954 min,该峰经二极管阵列光谱检测为单一组分。【结论】利用最佳的高速逆流色谱条件和高效液相色谱条件对小孢拟盘多毛孢Pestalotiopsismicrospora PM-1菌株代谢产物进行分离,可获得除草活性化合物——馏分A。
【Objective】 The purpose of this study was to establish the first separation of the herbicidal substances from the metabolites of Pestalotiopsis microspora PM-1 by high-speed counter-current chromatography (HSCCC). 【Method】 The optimum two-phase solvent system was n-hexane: ethyl acetate: methanol: water (4: 5: 4: 5, V / V / V / V) The lower phase (organic phase) as the mobile phase, normal phase elution. 【Result】 Four fractions were obtained under the conditions of flow rate of 2 mL / min, rotation speed of 900 r / min and detection wavelength of 254 nm. Among them, fraction Ⅱ showed strong activity on crabgrass. Fraction II was further separated by C18 column with high performance liquid chromatography (HPLC) using acetonitrile: water = 75:25 (V / V) as the mobile phase. Inhibition. The retention time of 7.954 min, the peak detected by diode array as a single component. 【Conclusion】 The herbicidal active compound-fraction A can be obtained from the metabolites of Pestalotiopsismicrospora PM-1 strain by using the best conditions of high-speed countercurrent chromatography and high performance liquid chromatography.