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(urate: oxygen oxidoreductase; EC

Uricase or Urate oxidase catalyzes the following reaction in all mammals except the primates and a few other species.

Uric Acid + 2H2O +O2 ------> Allantoin + H2O2 + CO2 
The enzyme is widely distributed in the tissues of lower mammals and in some microorganisms. Uricase is extensively used for the determination of uric acid in biological fluids. (Domagk, G.F. and Schlicke, H.H., Anal. Biochem., 22, 219, 1968).

The amount of enzyme which catalyzes the conversion of one micromole uric acid to allantoin per minute at 25°C.

The rate of oxidation of uric acid to allantoin is followed spectrophotometrically at 293 nm (Mahler, H.R. in The Enzymes, Boyer, P.D.; Lardy, H.A. and Myrback, K., eds., 8, 285, 1963, Academic Press, New York).

  1. 0.1 M Sodium borate buffer, pH 8.5.
  2. Uric acid (substrate). Dissolve 2 mg uric acid in 30 ml borate buffer. Adjust pH to 8.5 with KOH and dilute with buffer to give an absorbance of 0.50 at 293 nm.
  3. Uricase (enzyme) solution. Dissolve and then dilute in borate buffer to give a final concentration of 0.01-0.05 U/ml. Must be prepared fresh prior to assay.

  1. Set spectrophotometer (equipped with strip chart recorder and temperature control) at 293 nm and 25°C.
  2. Oxygenate the uric acid (substrate) solution for 5 minutes prior to use, then pipette 2.8 ml uric acid into a quartz cuvette.
  3. Incubate cuvette in the spectrophotometer at 25°C for 5 minutes to achieve temperature equilibration and then record absorbance at 293 nm (blank).
  4. Initiate the reaction by adding 0.2 ml Uricase (enzyme) solution to the cuvette. Record the decrease in absorbance at 293 nm for 6-8 minutes.
  5. Calculate µE293 nm/min

Activity (U/mg) = (ΔE293nm/min)(Total Vol.)(Enz. Diln.)
(12.3)(Enz. Vol.)(mg Enz./ml)

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