Water-soluble cationic boronate probe based on coumarin imidazolium scaffold: Synthesis, characterization, and application to cellular peroxynitrite detection

Aleksandra Grzelakowska; Julia Modrzejewska; Jolanta Kolińska; Marcin Szala; Monika Zielonka; Karolina Dębowska; Małgorzata Zakłos-Szyda; Adam Sikora; Jacek Zielonka; Radosław Podsiadły

doi:10.1016/j.freeradbiomed.2021.12.260

Water-soluble cationic boronate probe based on coumarin imidazolium scaffold: Synthesis, characterization, and application to cellular peroxynitrite detection

Free Radic Biol Med. 2022 Feb 1:179:34-46. doi: 10.1016/j.freeradbiomed.2021.12.260. Epub 2021 Dec 17.

Affiliations

¹ Institute of Polymer and Dye Technology, Faculty of Chemistry, Lodz University of Technology, Stefanowskiego 16, 90-537, Lodz, Poland. Electronic address: aleksandra.grzelakowska@p.lodz.pl.
² Institute of Polymer and Dye Technology, Faculty of Chemistry, Lodz University of Technology, Stefanowskiego 16, 90-537, Lodz, Poland. Electronic address: julia.modrzejewska@dokt.p.lodz.pl.
³ Institute of Polymer and Dye Technology, Faculty of Chemistry, Lodz University of Technology, Stefanowskiego 16, 90-537, Lodz, Poland. Electronic address: jolanta.kolinska@p.lodz.pl.
⁴ Institute of Polymer and Dye Technology, Faculty of Chemistry, Lodz University of Technology, Stefanowskiego 16, 90-537, Lodz, Poland. Electronic address: marcin.szala@p.lodz.pl.
⁵ Department of Biophysics, Medical College of Wisconsin, 8701 Watertown Plank Road, Milwaukee, WI, 53226, United States. Electronic address: mzielonka@mcw.edu.
⁶ Institute of Applied Radiation Chemistry, Faculty of Chemistry, Lodz University of Technology, Żeromskiego 116, 90-924, Lodz, Poland. Electronic address: karolina.debowska@p.lodz.pl.
⁷ Institute of Molecular and Industrial Biotechnology, Faculty of Biotechnology and Food Sciences, Lodz University of Technology, Stefanowskiego 2/22, 90-537, Lodz, Poland. Electronic address: malgorzata.zaklos-szyda@p.lodz.pl.
⁸ Institute of Applied Radiation Chemistry, Faculty of Chemistry, Lodz University of Technology, Żeromskiego 116, 90-924, Lodz, Poland. Electronic address: adam.sikora@p.lodz.pl.
⁹ Department of Biophysics, Medical College of Wisconsin, 8701 Watertown Plank Road, Milwaukee, WI, 53226, United States. Electronic address: jzielonk@mcw.edu.
¹⁰ Institute of Polymer and Dye Technology, Faculty of Chemistry, Lodz University of Technology, Stefanowskiego 16, 90-537, Lodz, Poland. Electronic address: radoslaw.podsiadly@p.lodz.pl.

PMID: 34923103
DOI: 10.1016/j.freeradbiomed.2021.12.260

Abstract

Peroxynitrite (ONOO^-) has been implicated in numerous pathologies associated with an inflammatory component, but its selective and sensitive detection in biological settings remains a challenge. Here, the development of a new water-soluble and cationic boronate probe based on a coumarin-imidazolium scaffold (CI-Bz-BA) for the fluorescent detection of ONOO^- in cells is reported. The chemical reactivity of the CI-Bz-BA probe toward selected oxidants known to react with the boronate moiety was characterized, and the suitability of the probe for the direct detection of ONOO^- in cell-free and cellular system is reported. Oxidation of the probe results in the formation of the primary hydroxybenzyl product (CI-Bz-OH), followed by the spontaneous elimination of the quinone methide moiety to produce the secondary phenol (CI-OH), which is accompanied by a red shift in the fluorescence emission band from 405 nm to 481 nm. CI-Bz-BA reacts with ONOO^- stoichiometrically with a rate constant of ∼1 × 10⁶ M^-1s^-1 to form, in addition to the major phenolic product CI-OH, the minor nitrated product CI-Bz-NO₂, which is not formed by other oxidants tested or via myeloperoxidase-catalyzed oxidation/nitration. Both CI-OH and CI-Bz-NO₂ products were also formed in the presence of cogenerated fluxes of nitric oxide and superoxide radical anion produced during decomposition of a SIN-1 donor. Using RAW 264.7 cells, we demonstrate the ability of the probe to report endogenously produced ONOO^-via fluorescence measurements, including plate reader real time monitoring and two-photon fluorescence imaging. Liquid chromatography/mass spectrometry analyses of cell extracts and media confirmed the formation of both CI-OH and CI-Bz-NO₂ in macrophages activated to produce ONOO^-. We propose the use of a combination of real-time monitoring of probe oxidation using fluorimetry and fluorescence microscopy with liquid chromatography/mass spectrometry-based product identification for rigorous detection and quantitative analyses of ONOO^- in biological systems.

Keywords: Coumarin-based probe; Mitochondria-targeted two-photon fluorescent probe; Peroxynitrite; Redox probe; Water-soluble fluorescent probe.

Publication types

Research Support, Non-U.S. Gov't

MeSH terms

Coumarins
Fluorescent Dyes
Oxidation-Reduction
Peroxynitrous Acid*
Superoxides
Water*

Substances

Coumarins
Fluorescent Dyes
Water
Superoxides
Peroxynitrous Acid