流动相的离子强度对RP-HPLC中抗生素色谱峰形的影响

在反相高效液相色谱(RP-HPLC)过程中,流动相的离子强度可显著影响可解离的化合物如抗生素的色谱峰形,原因在于流动相的离子强度较低时,色谱填料的固定相易于过载,使离子化的化合物的色谱峰形呈典型的过载特征——峰形接近直角三角形。随着进样量的增加,色谱区带中高浓度的前半部的保留时间减少,但色谱区带均在同一时刻结束,色谱峰的拖尾因子及峰宽均显著增加,同时被测物与有关物质间的分离也随之变差。选择合适的缓冲盐并适当地增加流动相的离子强度即可显著改善抗生素的色谱峰形,色谱峰的拖尾因子及峰宽均随之降低,同时被测物与有关物质间也更加易于分离。

Influence of the ionic strength of mobile phase on peak shape of antibiotics in RP-HPLC

The ionic strength of mobile phase can significantly affect the peak shape of ionogenic compounds such as antibiotics in reversed-phase high performance liquid chromatography(RP-HPLC).The influence may be attributed to overloading of ionogenic analytes in lower ionic strength mobile phase.In such phase,peak becomes increasingly right-angled triangle in shape with increasing sample load together with increasing peak width and tailing.The retention time of the high-concentration front decreases with increasing sample load,while the end of the peak tail has a constant retention time,equal to the symmetrical analytical peak.Due to considerably worse peak shapes,poorer resolution between the main component and its related substances might be observed.With the optimal buffer and the increase of ionic strength,significant improvement in peak shape of antibiotics could be achieved and consequently a decrease in the tailing factor,an increase in the apparent column efficiency as well as an efficient resolution were obtained.