Fluorescent Derivatization of Xanthurenic Acid and Nicotinic Acid with Horseradish Peroxidase in the Presence of Excess Hydrogen Peroxide

The fluorescent derivatization of tryptophan metabolites (xanthurenic acid, nicotinic acid, picolinic acid, and 3-hydroxyanthranilic acid) by the catalytic activity of horseradish peroxidase (HRP) was investigated in the presence of excess H2O2. Non-fluorescent xanthurenic acid (XA) and nicotinic acid (NA) were converted into a fluorescent compound with maximum excitation and emission wavelengths at 325 and 425 nm, and 318 and 380 nm, respectively. This fluorescent derivatization was developed for the fluorometric determination of trace amounts of XA and NA. The calibration curves were linear from 1.0 to 10.0 nmol XA and from 5.0 to 20.0 nmol NA in a 1.0-mL sample solution. The UV spectra of the reaction solutions suggested that compound III as an intermediate of HRP played an essential role in this fluorescent derivatization with HRP.