| Summary: | Derivatives of coumarin, containing oxidant-sensitive boronate group, were recently developed for fluorescent detection of inflammatory oxidants. Here, we report the synthesis and the characterization of 3-(2-benzothiazolyl)-7-coumarin boronic acid pinacol ester (<b>BC-BE</b>) as a fluorescent probe for the detection of peroxynitrite (ONOO<sup>–</sup>), with high stability and a fast response time. The <b>BC-BE</b> probe hydrolyzes in phosphate buffer to 3-(2-benzothiazolyl)-7-coumarin boronic acid (<b>BC-BA</b>) which is stable in the solution even after a prolonged incubation time (24 h). <b>BC-BA</b> is slowly oxidized by H<sub>2</sub>O<sub>2</sub> to form the phenolic product, 3-benzothiazol-2-yl-7-hydroxy-chromen-2-one (<b>BC-OH</b>). On the other hand, the <b>BC-BA</b> probe reacts rapidly with ONOO<sup>−</sup>. The ability of the <b>BC-BA</b> probe to detect ONOO<sup>–</sup> was measured using both authentic ONOO<sup>–</sup> and the system co-generating steady-state fluxes of O<sub>2</sub><sup>•</sup><sup>–</sup> and <sup>•</sup>NO. <b>BC-BA</b> is oxidized by ONOO<sup>–</sup> to <b>BC-OH</b>. However, in this reaction 3-benzothiazol-2-yl-chromen-2-one (<b>BC-H</b>) is formed in the minor pathway, as a peroxynitrite-specific product. <b>BC-OH</b> is also formed in the reaction of <b>BC-BA</b> with HOCl, and subsequent reaction of <b>BC-OH</b> with HOCl leads to the formation of a chlorinated phenolic product, which could be used as a specific product for HOCl. We conclude that <b>BC-BA</b> shows potential as an improved fluorescent probe for the detection of peroxynitrite and hypochlorite in biological settings. Complementation of the fluorescence measurements by HPLC-based identification of oxidant-specific products will help to identify the oxidants detected.
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