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

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 x 10(-8) mol L-1 of peroxynitrite. A liner calibration graph is obtained over the peroxynitrite concentration range 5.60 x 10(-7) to 2.10 x 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. (C) 2004 Elsevier B.V. All rights reserved.