Enzymatic method for assaying uric acid in serum with a new tetrazolium salt produces water-soluble formazan dye

Objectives: To apply an enzymatic method for assaying uric acid in serum based on the uricase (EC 1.7.3.3)-catalase (EC 1.11.1.6)formaldehyde dehydrogenase (FADH, EC 1.2.1.46) coupled with the new tetrazolium salt producing a water-soluble formazan dye as an indicator system. Unlike the traditional tetrazolium salts, e.g., iodonitrotetrazolium (INT) and nitrotetrazolium blue (NTB), the corresponding formazan dye produced did not absorb to the test tube and the reaction cells of the analyzer. Moreover, this formazan dye had a higher water solubility and more sensitivity. Design and Methods: A two electron reduction of tetrazolium salt with NADH, produced by the uricase-catalase-FADH reaction, was mediated by an electron carrier, 1-methoxy PMS. The generation of formazan, monitored at 440 nm, was proportional to the concentration of uric acid in serum. The sensitivity of this method was about 1.5 times that of the peroxidase-TOOS [N-ethyl-N-(2- hydroxy-3-sulfopropyl)-m-toluidine] method. The assay was evaluated with TBA- 80FR. NEO biochemical analyzer. The average within-run and between-day imprecision (CV) were 0.75-2.44% and 3.20-3.33%, respectively. Results: Results of the proposed method (y) correlated well with those determined by the conventional chromogen method (x): y = 0.970 x - 0.018 mmol/L (Sy l x = 0.019 mmol/L, r = 0.993, n = 67). Conclusions: We also present data showing that the method is rapid, relatively free of interference, and amenable to automation.