On-the-fly fluorescence lifetime detection in HPLC using a multiharmonic Fourier transform phase-modulation spectrofluorometer

Frequency-domain fluorescence lifetime detection has been demonstrated recently as a technique for detecting and resolving overlapping peaks on-the-fly in reversed-phase high-performance liquid chromatograghy (RP-HPLC). However, instrument limitations necessitate multiple injections of the same sample for peak resolution. The introduction of a commercially available multiharmonic Fourier transform spectrofluorometer (MHF) eliminates this problem and offers other advantages as well. The MHF data acquired onthe-fly during a single chromatograghic run contain all of the multifrequency information needed to determine lifetimes and to indicate and resolve overlapping peaks at intervals as short as several milliseconds