A sensitive method for the simultaneous determination of urinary imidazoleacetic acid and N tau- and N pi-methylimidazoleacetic acids which employs high-performance liquid chromatography with fluorescence detection is described. The compounds were converted into the corresponding fluorescent esters by reaction with 4-bromomethyl-7-methoxycoumarin. These derivatives are separated by liquid chromatography on a Radial-Pak silica column. The detection limits for imidazoleacetic acid and N tau-and N pi-methylimidazoleacetic acids in urine are 15, 10 and 20 pmol/ml, respectively. The 24-h urinary excretion of imidazoleacetic acid and N tau-and N N pi-methylimidazoleacetic acids by healthy persons was 5.7-39.9, 4.3-24.6 and 1.5-19.3 nmol/mg of creatinine, respectively.