Kinetic characterisation and thermal inactivation study of red alga (Mastocarpus stellatus) peroxidase

Abstract

Peroxidase (POD) was extracted from red alga (Mastocarpus stellatus) using Triton X-114 and characterised by UV-spectrophotometry. Optimum activity using 2,2´ -azinobis(3-ethylbenzothiazolinesulphonic acid) (ABTS) as the H-donor was obtained at pH 5.0. In the presence of the anionic detergent, sodium dodecyl sulphate (SDS), however, POD was inactivated at all the pH values studied and totally inactivated at 1 mM SDS. When the enzyme was kinetically characterised, the KM and Vm values for ABTS were found to be 13 mM and 40 lM/min, respectively. In addition, when the H2O2 concentration was increased, at a fixed concentration of ABTS, the activity was inhibited at the highest H2O2 concentrations. In a study of the effect of several reducing agents, L-cysteine was found to be the most active. A thermal inactivation study showed a first-order inactivation kinetic, and the Arrhenius plot yielded a straight line with a slope equivalent to an activation energy of 121.6 kJ/mol. Significant inactivation occurred at temperatures of >35 C, with >90% of the relative activity being lost after only 5 min of incubation at 48.4 C.