Catalase Assay

There are numerous assays for catalase. Gregory and Fridovich (1974) report on a sensitive activity stain for catalase applicable to a polyacrylamide gel electrophoretogram, Haining and Legan (1972) describe a polarographic assay utilizable in tissue homogenates, and Kroll et al. (1989) discuss a rapid method for estimating the bacterial content of foods. The subject has been reviewed by Maehly and Chance (1954) and Chance and Maehly (1955). The assay used at Worthington follows:


Method: Essentially that described by Beers and Sizer (1952) in which the disappearance of peroxide is followed spectrophotometrically at 240 nm. One Unit decomposes one micromole of H2O2 per minute at 25°C and pH 7.0 under the specified conditions.


  • 0.05 M Potassium phosphate, pH 7.0
  • 0.059 M Hydrogen peroxide (30%) in 0.05 M potassium phosphate, pH 7.0


Immediately prior to use dilute the enzyme in 0.05 M phosphate buffer, pH 7.0 to obtain a rate of 0.03-0.07 ΔA/min.



Adjust the spectrophotometer to 240 nm and 25°C.

Pipette into each cuvette as follows:

Reagent grade water 1.9 ml
0.059 M Hydrogen peroxide 1.0 ml

Incubate in spectrophotometer for 4-5 minutes to achieve temperature equilibration and to establish blank rate if any. Add 0.1 ml of diluted enzyme and record decrease in absorbance at 240 nm for 2-3 minutes. Calculate ΔA240/min from the initial (45 second) linear portion of the curve.



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