Direct Examination of Elution Nonequilibrium in Liquid Chromatography by Laser-Induced Fluorescence

Abstract Nonequilibrium interactions, which are known to influence broadening throughout the column, become very important at the exit of the chromatographic column. Theoretical predictions, derived herein, show a substantial increase in length variance and decrease in concentration when the solute is detected off-column. The discrepancy between on-and off-column values for variance and concentration is predicted to increase markedly with capacity factor. Experimental investigation of elution nonequilibrium is accomplished by measuring the length variance and concentation for a homologous series of model solutes both on-and off-column by laser-induced fluorescence. Excellent agreement of experimental measurement with theoretical predictions is seen for solutes with capacity factors ranging from one to nine. These results have interesting implications for the general elution problem in chromatographic separations.