Determination of paracetamol (acetaminophen) by HPLC with post-column enzymatic derivatization and fluorescence detection

A novel method is described for the determination of paracetamol (acetamino phen; N-acetyl-4-aminophenol) in urine. After reversed-phase HPLC separatio n, paracetamol is oxidized by H2O2 with horseradish peroxidase catalysis. D etection is performed fluorimetrically at an excitation wavelength of 329 n m and an emission wavelength of 435 nm. Urine samples were spiked with para cetamol, diluted, and injected directly without further pretreatment. Under these conditions, the limit of detection was 2 x 10(-8) mol L-1, and the l imit of quantification was 7 x 10(-8) mol L-1. The method was validated by two different approaches based on HPLC with UV-Vis detection.