A simple uric acid assay by using 3‐hydroxytyramine as a chromogenic colorimetric sensor in human serum samples: Density functional theory supported mechanistic approach

Abstract

A simple, fast, sensitive, convenient, and economical accurate enzymatic colorimetric sensor is developed for the quantification of the clinically important biomarker uric acid (UA) in human serum samples. The method is based on the quantification of UA using 3-hydroxytyramine (3-HT) as a chromogenic substrate by using peroxidase (POD) and uricase (UOx). UOx catalyzes the oxidation of UA where H2O2 is generated in-situ to produce allantoin and CO2. The POD in oxidative coupling of peroxide in presence of 3-HT, forms an intense orange-colored product with λmax = 500 nm. The reaction was carried out in a citric acid-tripotassium-citrate buffer (12.5 mM) of pH = 6.8 at room temperature. Computational calculations were done by Gaussian-16 employing the B3LYP functional basis set. The standard curve for UA was found to be linear between 12.3–297.3 and 3.07–396.5 μM by rate and fixed time method, respectively. The LOD and LOQ for UA were 1.5 and 2.9 μM, respectively. Within-day and day-to-day precision was 1.50–3.07 and 3.10–4.16% (n = 7), respectively. The accuracy ranges for UA concentrations of 24.7, 198 and 297.3 μM were 87%–101.50%, 90%–105%, and 89%–107%, respectively. The UA recovery range by the proposed method was 98.00%–100.47% with a mean recovery of 99.5% and has A good correlation coefficient of 0.981 with the standard method.