CAT activity was measured spectrophotometrically by measuring the rate of
H2O2 disappearance at 240 nm (∆ε = 43.6 mM-1 cm-1) according to the method
Patterson et al. (1984). The reaction mixture contained 50 mM sodium phosphate
buffer (pH 7.0), 19 mM H2O2 and 25 µg protein in a final volume of 3 ml. The activity was determined by the oxidation of H2O2 at 240 nm.
how can i calculate my results using this formula (∆ε = 43.6 mM-1 cm-1)
Hi Benedict,
If you have a calibration curve, you can use sth like this:
1. (((Asample-Ablank) - y-int)/slope)* total volume of your sample in well or cuvette/volume of protein or cell lysate = uM of sample
A-means absorbance
y-int and slop you will get from callibration curve.
2. (uM of sample/time of incubation with hydrogen peroxide)x sample dilution = nmol/min/ml
If you want you can standarize per mg of protein, you have to only divide by protein concentration.
I hope it will help you :)
If not, here you have another method:
https://www.bioclot.com/PDF/catalase_kinetic_assay_v.05.pdf
Mike
Hi Benedict,
CAT activity was determined as decrease in hydrogen peroxide. The enzymatic reaction of H2O2 degradated by CAT was terminated by the addition of ammonium molybdate, and the rest H2O2 combined with ammonium molybdate generated a light-yellow complex that could be measured spectrophotometrically at 405 nm.
Please refer to the PDF This should solve all our problems. All the best.
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