Two-Photon Excitation of Fluorogenic Probes for Redox Metabolism:Dramatic Enhancement of Optical Contrast Ratio by Two-Photon Excitation

摘要

Fluorogenic probes for redox metabolism are shown to exhibit two-photon absorption using femtosecond pulses from a Ti/sapphire laser.The probes consist of a coumarin or naphthalene core and a ketone-alcohol functional group.The probe design is based on the changes in fluorescence properties and two-photon cross section values when the ketone derivative is reduced to the corresponding alcohol.The resulting contrast ratio of the fluorescence differs significantly from that obtained by one-photon excitation.This phenomenon was demonstrated with all three switches examined herein and represents an attractive approach to modulation of emission properties of molecular switches.The practical applicability of the two-photon-excitation redox switch was demonstrated in an enzyme-catalyzed transformation.The nonfluorescent probe 1 is efficiently converted by AKR1C3,a human 3a-hydroxysteroid dehydrogenase,to the fluorescent derivative 2.Using two-photon excitation(775 nm),we achieved a large contrast ratio between the fluorescence in the presence and the absence of the enzyme of ~300.