You will have to provide more details of your method.

I am guessing that you are using an absorbance measurement, wherein the catalase activity is measured on the basis of the change in absorbance. If this is correct, you have  a few options for converting this absorbance change into a measurement of enzyme activity. Can you clearly state the goal of the measurement? Do you want to measure the specific activity of catalase in a sample, for example?

Hi, Adam...

Yes, I am working with the change in absorbance, using the specifications to microplate reader described by this autor (http://www.ncbi.nlm.nih.gov/pmc/articles/PMC2062561/) but i hadn't found any equations that describe the calculations to microplate reader, only assays with cuvette.

i want measure the specific activity of catalase in a sample, how is possible this?

The only difference between the calculations is that the pathlength in the cuvette is 1 cm, but the pathlength in the microplate depends on the volume in the well. The pathlength is the distance that light passes through the solution. Pathlength is needed to use Beer's Law to convert absorbance to concentration. However, it is not really necessary to know the pathlength to make this conversion if you prepare a standard curve of product, using the same exact volume as in the assay.

Specific activity is usually defined as micromoles of product formed per minute per mg of  enzyme, with the exact conditions of the assay stated. So you need to measure the rate of product formation and the amount of enzyme that is put into the reaction. You get the rate of product formation by measuring the slope of the absorbance versus time curve (at the start of the reaction - the initial rate) and apply the standard curve to convert the absorbance change to product concentration.

Hi Diego, 

Did you figured out a formula for calculating enzyme activity using plate reader? I am also using microtiter plate for measuring absorbance changes for some enzymes!

Thanks 

For Microplate calculations ..pathlength should be considered 0.5 cm

hey please tell me how to calculate enzyme activity using plate reader and what is the significance of taking absorption at a particular difference of time for the fix interval ?

for ex. i am taking absorption at 405 nm at an interval of 30 sec. for 10 min.

and then how we will calculate the change in absorbance per unit time from this ?

You plot a graph of absorbance versus time. The slope of the linear initial part of the graph is the change in absorbance per unit of time.

If you make a calibration curve of absorbance versus product concentration, you can convert change in absorbance per unit time into change in product concentration per unit time, which is the rate of the reaction.