Cheletropic traps for the fluorescence spectroscopic detection of nitric oxide (nitrogen monoxide) in biological systems

The concept of nitric oxide cheletropic traps (NOCT), which relies on the E SR spectroscopic detection of nitroxide radicals formed on trapping of NO b y the NOCTs, has been extended to NOCTs that incorporate a fluorophoric sys tem and form a new fluorophore on reaction with NO (FNOCT = fluorescent NOC T). The synthesis of the acetoxymethylesters of two FNOCTs, which can be lo aded into cells, is described. Inside the cells the esters are enzymaticall y hydrolysed, thus allowing intracellular trapping of NO. The primary nitro xides formed on reaction with NO are reduced to the corresponding hydroxyla mines under physiological conditions, thus remedying the fluorescence-quenc hing properties of nitroxide radicals. It is demonstrated that NO formed by lipopolysaccharide(LPS)-activated alveolar macrophages can easily be detec ted by these FNOCTs on a single cell basis.