Fluorescence Quenching Effect of a Highly Active Nitroxyl Radical on 7-amino-4-methylcoumarin and Glutathione Sensing

Nitroxyl radical compounds, such as 2,2,6,6-tetramethylpiperidine 1-oxyl (TEMPO), are stable radical compounds with a variety of unique characteristics, including fluorescence quenching. In this study, we investigated the fluorescence quenching effect of nortropine N-oxyl (NNO), which is a highly active nitroxyl radical that is more active than TEMPO in oxidation catalysis. The fluorescence intensity of 7-amino-4-methylcoumarin (AMC) was quenched by NNO and TEMPO to 5% and 95% of the initial fluorescence intensity, respectively, indicating highly efficient quenching by NNO. In addition, we used this reaction to measure glutathione concentration. The quenching effect of NNO was abrogated by the chemical reaction with glutathione, resulting in restoration of AMC fluorescence. This response was observed at glutathione concentrations from 10 µM to 1 mM, and good calibration curves were obtained from 10 to 250 µM.