A Direct Evidence of Stoichiometric Displacement between Insulin and Methanol in Reversed Phase Liquid Chromatography

Ｗith four kinds of mobile phases,methanol-water,ethanol-water,2-propanol and acetonitrile-water(all containing 0.1％rifluroacetic acid),the displacement between solute and solvent in RPLC was proved to be universal in frontal analysls(FA).Based on the measured Z value in usual RPLC to be a constant and the quantitative determination of methanol increment in mobile phase in FA,the stoichiometric displacement(SD)between insulin and methanol was directly proved by the experiment.The SD was also proved to occur only on about the one-fourth of the maximum amount of adsorbed methanol in the bonded phase layer(BPL)without any dynamic problem of mass transfer,while in FA,the SD firstly occurs on the surface of the BPL and then gradually sinks into the deeper sites companied with a dynamic problem.Although the displaces soplvent by the same solute is less in the former case,the SD is independent of how deep of the solute enters the BPL.In addition,the adsorbed amount of solute on and adsorbent not only depends on the numbers of the adsorbed layer on the adsorbent surface,but also on the extent of the complete removal of the displace able solvent in the BPL.The pyhsical fundamental of the SD and the methodoloby for investigation were also discussed.

A Direct Evidence of Stoichiometric Displacement between Insulin and Methanol in Reversed Phase Liquid Chromatography

With four kinds of mobile phases, methanol water, ethanol water, 2 propanol and acetonitrile water (all containing 0 1% triflu roacetic acid), the displacement between solute and solvent in RPLC was proved to be universal in frontal analysis (FA). Based on the measured Z value in usual RPLC to be a constant and the quantitative determination of methanol increment in mobile phase in FA, the stoichiometric displacement (SD) between insulin and methanol was directly proved by the experiment. The SD was also proved to occur only on about the one fourth of the maximum amount of adsorbed methanol in the bonded phase layer (BPL) without any dynamic problem of mass transfer, while in FA, the SD firstly occurs on the surface of the BPL and then gradually sinks into the deeper sites companied with a dynamic problem. Although the displaced solvent by the same solute is less in the former case, the SD is independent of how deep of the solute enters the BPL. In addition, the adsorbed amount of solute on an adsorbent not only depends on the numbers of the adsorbed layer on the adsorbent surface, but also on the extent of the complete removal of the displaceable solvent in the BPL. The physical fundamental of the SD and the methodology for investigation were also discussed.