Uric acid determination using uricase and the autotransducer molecular absorption properties of peroxidase

Uric acid (UA) is determined using the UV-vis molecular absorption properties of peroxidase (HRP). The method as a whole involves UA oxidation in the presence of uricase (UOx), giving H₂O_2. The H₂O₂ then reacts with HRP forming the compound I species which returns to its initial form by reaction with UA and intramolecular reduction. The molecular absorption changes of HRP at 420 nm during the reaction enable the UA to be determined. A mathematical model relating the analytical signal to UA, UOx and HRP has been developed and experimentally validated. The possibility of carrying out both enzymatic reactions sequentially or simultaneously is discussed, the latter option producing better analytical performances. The method permits UA determination in the range 1.5 × 10⁻⁶-4.0 × 10⁻⁵ M, with an R.S.D. of about 3% (n = 5, 1.5 × 10⁻⁶ M UA). It has been applied to analyte determination in synthetic serum samples.