A novel method for determination of peroxynitrite based on hemoglobin catalyzed reaction Original Research Article

A novel spectrofluorimetric method for the determination of peroxynitrite is proposed. The method is based on a mimetic enzyme catalyzed reaction with hemoglobin as the catalyst and l-tyrosine as the substrate. A new fluorescent substance is produced that might probably be the coupled dimmer of tyrosine, which, instead of nitryl-tyrosine, is likely to be a new marking substance of ONOO− injury in vivo. Kinetics of the reaction is studied and the possible reaction mechanism is also recommended. The proposed method is simple and highly sensitive with a detection limit of 5.00 × 10−8 mol L−1 of peroxynitrite. A liner calibration graph is obtained over the peroxynitrite concentration range 5.60 × 10−7 to 2.10 × 10−5 mol L−1, with a correlation coefficient of 0.9983. Interferences from some amino acids and metal ions normally seen in biological samples, and also some anions structurally similar to ONOO− are studied.