Revisiting the Chemistry and Photophysics of 3-(N-Methylpyridinium-4-yl)Coumarins for Designing "Covalent-Assembly" and "Molecular Disassembly" Fluorescent Probes

The constant need for high-performance aniline- or phenol-based fluorophores suitable for the construction of activity-based fluorescent probes, led us to study both synthesis and photophysics of under-explored C3-N-methylpyridinium-4-yl substituted 7-(dialkylamino)/7-hydroxycoumarins. Indeed, in the field of photoactive organic molecules, the positively charged N-alkylpyridinium-4-yl groups are often used as acceptor units to dramatically impact spectral features through promoting intramolecular charge transfer (ICT) process. They are also known as effective water-solubilizing and cell organelle (mitochondria) targeting moieties. The poor fluorescence efficiency of cationic 7-hydroxycoumarin derivatives in aqueous physiological conditions was highlighted and rationalized by the predominance of a neutral quinonoid form in such buffer medium. The ability of the excited singlet state (S1) of this neutral species to undergo intersystem crossing (ISC) to triplet state (T1) was partly supported by phosphorescence measurements of singlet oxygen (1O2). We took advantage of superior green-emissive properties of 7-(diethylamino)-3-(N-methylpyridinium-4-yl)coumarin to successfully design and validate a novel small-molecule fluorescent probe for the detection of alkaline phosphatase (ALP), based on the "covalent-assembly" principle. In last part of this study, a practical use of ortho-formylated 7-hydroxy-3-(N-methylpyridinium-4-yl)coumarin was considered with the synthesis of a Fe(III)-salen complex known to act as effective "molecular disassembly" probe for fluorogenic sensing pyrophosphate (PPi) ions in aqueous media, and through the promising approach recently proposed by the Zelder group (Chimia 2020, 74, 252-256).