The fluorescence upconversion technique has been applied to examine the picosecond fluorescence decay kinetics of flavin adenine dinucleotide (FAD) in aqueous solution. In the observation range of 30 ps three fluorescent lifetimes can be distinguished. The shortest-lived component (similar to1 ps) arises from water relaxation around the excited flavin. The 9-ps component originates from the intramolecular complex between flavin and adenine, whereas the nanosecond decay is attributed to the unstacked form of FAD. The spectra of the three forms are derived from global analysis of decay curves at different emission wavelengths and time regimes using a triple exponential function. It is assumed that the amplitude belonging to the nanosecond fluorescence component reflects the steady-state fluorescence spectrum. Fluorescence anisotropy to its maximum value of 0.4 is instantaneously created. (C) 2003 Elsevier B.V. All rights reserved.