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目的 筛选分离纯化卟啉锰 TFO 吖啶化合物的简单有效的方法。方法 以固相亚磷酰胺法合成吖啶、6碳氨基连接臂修饰的TFO ,并以温和法切割和脱保护 ;四苯基卟啉四羧酸经金属化、酯化后 ,与上述TFO偶联 ;分别以变性聚丙酰胺凝胶电泳、寡核苷酸纯化柱、薄层色谱法分离纯化偶联反应产物。结果 以凝胶电泳分离偶联反应产物时 ,偶联反应产物带、卟啉锰活性酯带位于同一水平 ,未分离出独立的卟啉锰 TFO 吖啶化合物带。以寡核苷酸纯化柱分离偶联反应产物时 ,紫外分光光度仪检测结果显示 ,回收物仍为卟啉锰 TFO 吖啶化合物与游离的卟啉锰活性酯的混合物。以薄层层析分离偶联反应产物时 ,在卟啉锰活性单酯带后 ,出现一条绿色月牙型带 ,紫外光光谱分析显示 ,此带回收物同时具有寡核苷酸和卟啉锰的特征吸收峰 (UVλmax =2 6 0、4 6 8nm) ,质谱分析结果证实 ,回收物分子量实测值与卟啉锰 TFO 吖啶化合物理论一致。结论 薄层色谱法可简单有效分离纯化卟啉锰 TFO 吖啶化合物。
Objective To screen a simple and effective method for the isolation and purification of TFO acridine compounds. Methods The acridine and 6-carbon amino linker modified TFO was synthesized by solid-phase phosphoramidite method and cleaved and deprotected by mild method. The tetraphenylporphyrin tetracarboxylic acid was metallized and esterified, Respectively; the denatured polyacrylamide gel electrophoresis, oligonucleotide purification column, TLC separation and purification of the coupling reaction product. Results When the coupling reaction product was separated by gel electrophoresis, the product of the coupling reaction and the active bands of the porphyrin manganese were at the same level, and no separate porphyrin manganese TFO acridinium band was isolated. When the coupling reaction product was separated and purified by an oligonucleotide purification column, the UV spectrophotometer test showed that the recovered product was still a mixture of the porphyrin manganese TFO acridine compound and the free active manganese porphyrin. When the coupling reaction product was separated by thin-layer chromatography, a green crescent-shaped band appeared after the active manganese monofunctional zone of porphyrin. The ultraviolet spectrum showed that this band recovered both oligonucleotide and porphyrin manganese The characteristic absorption peak (UV λmax = 2660, 466 nm) confirmed by mass spectrometry that the molecular weight of the recovered product was in agreement with the theory of the porphyrin manganese TFO acridine compound. Conclusion Thin layer chromatography can be used to separate and purify TFO acridine compounds easily and effectively.