一种通过反相高效液相色谱法快速可靠地分析核苷酸库的方法。

Rapid and reliable method for the analysis of nucleotide pools by reversed-phase high-performance liquid chromatography.

作者信息

Martinez-Valdez H, Kothari R M, Hershey H V, Taylor M W

出版信息

J Chromatogr. 1982 Oct 1;247(2):307-14. doi: 10.1016/s0021-9673(00)85954-3.

Abstract

A rapid and reliable protocol for the simultaneous separation of ribo-, deoxyribo- and cyclic nucleotides has been developed using high-performance liquid chromatography on a C18 microBondapak column and isocratic elution with ammonium phosphate buffer 0.2 m, pH 5.1). Resolution of deoxyribonucleotides has been confirmed by performing resolution before and after periodate oxidation. The general order of elution is ribonucleotides, deoxyribonucleotides and cyclic nucleotides. While periodate oxidation improved the clarity of separation of deoxyribonucleotides by eliminating ribonucleotides, incorporation of methanol in the eluent shortened the retention time of cyclic nucleotides. The application of this method to a complex biological system is reported.

摘要

已开发出一种快速可靠的方案，用于同时分离核糖核苷酸、脱氧核糖核苷酸和环核苷酸。该方案采用在C18微孔键合硅胶柱上进行高效液相色谱，并使用0.2m、pH 5.1的磷酸铵缓冲液进行等度洗脱。通过在高碘酸盐氧化前后进行分离，证实了脱氧核糖核苷酸的分离效果。洗脱的一般顺序是核糖核苷酸、脱氧核糖核苷酸和环核苷酸。虽然高碘酸盐氧化通过去除核糖核苷酸提高了脱氧核糖核苷酸分离的清晰度，但在洗脱液中加入甲醇缩短了环核苷酸的保留时间。本文报道了该方法在复杂生物体系中的应用。