剧毒卡罗藻GM5株中标志性毒素4,5-dihydro-KmTx2的色谱制备

Chromatographic preparation of 4,5-dihydro-KmTx2 from Karlodinium veneficum GM5 strain

建立了剧毒卡罗藻GM5株中标志性毒素4,5-dihydro-KmTx2的分离纯化体系,通过高效液相色谱-四极杆/静电场轨道阱高分辨质谱(UHPLC-Q-Orbitrap HRMS)对其纯化效果进行评价.过滤后的培养液经大型固相萃取柱(YMC-GEL ODS-A-HG,80 cm×10 cm,12 nm,S-50μm)进行富集,依次用体积分数为10%、40%和60%的甲醇溶液进行淋洗,体积分数为80%的甲醇溶液进行洗脱,得到目标化合物粗提液(纯度为51.3%).采用半制备型高效液相色谱技术进一步分离纯化,使得目标化合物纯度高达93.2%.该方法提取及纯化效率高,可操作性强,可以为4,5-dihydro-KmTx2的规模化生产及功能活性研究提供技术支持.

A separation and purification system for the toxin 4,5-dihydro-KmTx2 in Karlodinium veneficum GM5 strain at the end of the platform was established.The purification effect of the system was evaluated by high-performance liquid chromatography-quadrupole/electrostatic field orbitrap high-resolution mass spectrometry(UHPLC-Q-Orbitrap HRMS).First,the collected culture solution following the removal of the algae was enriched on a large solid-phase extraction column(YMC-GEL ODS-A-HG,10 cm×80 cm,12 nm,S-50μm).The results showed that leaching was accomplished in sequence by using 10%,40% and 60% methanol solution.The purity of the crude extract obtained by elution with 80%methanol was 51.3%.The crude extract was separated and purified by semi-preparative HPLC.The purity of the 4,5-dihydro-KmTx2 monomer was 93.2%with gradient elution using a methanol-water mixture as the mobile phase.The method has high extraction efficiency and operability,which can provide technical support for large-scale production and functional activity research of 4,5-dihydro-KmTx2.