In memoriam Hanns FischerThe synthesis and properties of the new fluorescent nitric oxide cheletropic trap (FNOCT) 14, designed for the trapping and quantification of nitric oxide (NO) production in chemical and biological systems, is described (Scheme 3). The dicarboxylic acid 14 and the corresponding bis[(acetyloxy)methyl] ester derivative 15 of the FNOCT contain a 2-methoxy-substituted phenanthrene group as fluorophoric unit. The fluorescence of the reduced NO adduct of this FNOCT (l exc 320 nm, l em 380 nm) is pH-independent. Trapping experiments were carried out in aqueous buffer solution at pH 7.4 with nitric oxide being added as a bolus as well as being released from the NO donor compound MAHMA NONOate (= (1Z)-1-{methyl[6-(methylammonio)hexyl]amino}diazen-1-ium-1,2-diolate), indicating a trapping efficiency of ca. 50%. In a biological application, nitric oxide was scavenged from a culture of lipopolysaccharide-stimulated rat alveolar macrophages. Under the applied conditions, a production of 11.1 AE 1.5 nmol of NO per hour and per 10 5 cells was estimated.Introduction. We have been concerned with the development of a reaction for the direct detection of NO which we discovered in 1992 [7 -10]. Although nitric oxide as a radical is rather unreactive towards organic molecules under normal conditions, it was found that it reacts readily with o-quinodimethanes (= 5,6-dimethylenecyclohexa-1,3-diene) to form cyclic nitroxide radicals in a kind of cheletropic reaction. This reaction is shown in Scheme 1 for the molecule first used for this purpose, tetramethyl-o-quinodimethane 1 [7]. EPR Spectroscopy seemed to be the method of choice for the detection of the thus formed nitroxide 2.The reactive o-quinodimethanes were named nitric oxide cheletropic traps (NOCTs). To apply this method to biological systems, a number of problems had to

The synthesis and the structural and spectroscopic characterization of nonfluorescent, pyrene-based cyclic o-quinodimethanes are reported. These compounds react efficiently with nitric oxide (NO) in a formal cheletropic manner, by which the fluorescent aromatic pyrene system is regenerated. The NO trapping capabilities and kinetics of the fluorescent nitric oxide cheletropic traps (FNOCTs) are assessed in THF and buffered aqueous solution by ESR, UV/Vis, and fluorescence spectroscopy, by employing NO solutions and NO released from N-diazeniumdiolates (NONOates). Prototypal biological applications include the quantitation of NO production from cultured rat alveolar macrophages and the endothelium of porcine aorta, which demonstrate a sensitivity for NO detection in the nanomolar range.

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