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Two problems are encountered in degassing hexane-based mobile phases. First, when fluorescence is used for the detection of vitamins A and E, the peak responses observed for consecutive injections of the same sample decrease over time due to quenching by oxygen dissolved in the mobile phase. Second, a rapid increase in retention time is observed with consecutive injections due to the preferential evaporation of the modifier, isopropanol (0.5%) in hexane. The rapid evaporation is due to the formation of an azeotrope between isopropanol and hexane with a boiling point of 63°C. A discussion of possible solutions to the two problems is given.

Chromatographic Problems Related to Mobile Phase Degassing and Fluorescence Detection