Structural Modifications of Nile Red Carbon Monoxide Fluorescent Probe: Sensing Mechanism and
                                           Applications

               Dominik Madea,^1) Marek Martínek,^1) Lucie Muchová,^2) Jiří Váňa,^3)

                                   Libor Vítek,^2) Petr Klán^1)

         ^1) Department of Chemistry and RECETOX, Faculty of Science, Masaryk University,

            Kamenice 5/A8, 625 00 Brno, Czech Republic, email: dominik.madea@seznam.cz

^2) Institute of Medical Biochemistry and Laboratory Diagnostics, 1^st Faculty of Medicine, Charles
  University, Na Bojišti 3, 121 08 Praha 2, Czech Republic^ 3) Institute of Organic Chemistry and
    Technology, Faculty of Chemical Technology, University of Pardubice, Studentská 573, 532 10
                                     Pardubice, Czech Republic


Carbon monoxide (CO) is a cell-signaling molecule (gasotransmitter)[1] produced endogenously by
oxidative catabolism of heme, and its spatial and temporal sensing at the cellular level is still
an open challenge.[2]

Synthesis, optical properties and study of the sensing mechanism of Nile-red Pd-based CO
chemosensors,[3] structurally modified by core and bridge substituents, in methanol and aqueous
solutions are reported in this work. The sensing fluorescence “off-on” response of
palladacycle-based sensors possessing low background fluorescence arises from their reaction with
CO to release the corresponding highly fluorescent Nile red derivatives in the final step. Our
kinetic study showed that electron-withdrawing and electron-donating core substituents affect a
rate-determining step of the reaction. More importantly, the substituents were found to have a
substantial effect on the Nile red sensor fluorescence quantum yields, hereby defining the sensing
detection limit.

The highest overall fluorescence and sensing rate enhancements were found for a 2-hydroxy
palladacycle derivative, which was used in subsequent biological studies on mouse hepatoma cells as
it easily crosses the cell membrane and qualitatively traces localization of CO within the
intracellular compartment with the linear quantitative response to increasing CO concentrations.


[1] Ryter, S. W.; Alam, J.; Choi, A. M. K. Physiol. Rev. 2006, 86, 583-650.

[2] Wu, L.; Wang, R. Pharmacol. Rev. 2005, 57, 585-630.

[3] Liu, K. Y.; Kong, X. Q.; Ma, Y. Y.; Lin, W. Y. Angew. Chem. Int. Ed. 2017, 56, 13489-13492.