A fluorescence method for the determination of photochemically generated peroxynitrite in seawater

Anal Chim Acta. 2020 Oct 2:1132:83-92. doi: 10.1016/j.aca.2020.06.063. Epub 2020 Jul 26.

Abstract

Peroxynitrite (ONOO-) is a reactive species which can degrade several classes of organic compounds via both oxidation and nitration reactions. Despite the fact that the photochemical precursors; superoxide (O2·-) and nitric oxide (NO·) radicals, have been measured in seawater under simulated solar radiation, there is no published report on actual measurements of photoformed peroxynitrite in seawater. Hence, this attempt was made to develop a fluorescence method by using coumarin boronic acid (CBA) as a chemical probe. CBA reacts with ONOO- to yield fluorescent 7-hydroxycoumarin (COH). COH was determined by reversed‒phase isocratic HPLC with fluorescence detection at excitation/emission wavelengths of 332/475 nm, respectively. COH standards calibration is linear in range of (0.25-100) × 10-9 M. The detection limit, defined as 3σ of the lowest ONOO- photo-formation rates obtained within 10- min irradiation, was 1.66 × 10-12 M s-1. This method is very precise as coefficient of variation for triplicate measurements of COH photo-formation rates was a maximum of 0.052. Experimental procedures were optimized to handle potential interference by hypochlorite, and the method was applied to measure ONOO- in 13 surface seawater samples obtained from the Seto Inland Sea, Japan. ONOO- photo-formation rates, steady‒state concentrations and lifetimes were determined to be (0.06-5.13) × 10-9 M s-1, (0.98-6.11) × 10-11 M and (0.01-0.16) s, respectively.

Keywords: Fluorescence; Liquid chromatography; Nitric oxide; Peroxynitrite; Photochemical; Superoxide.