Mango (Mangifera indica) belonging to Anacardiaceae family is a tropical fruit-bearing plants of Asian origin which grows well in tropical regions and it is considered as the king of fruits. Peroxidases (E.C.126.96.36.199) (POD) are a group of oxidoreductases that catalyse the reduction of peroxides, such as hydrogen peroxide and the oxidation of a variety of organic and inorganic compounds. POD activity has been identified in plants, microorganisms and animals where they play important roles. POD are used for clinical and industrial purposes but like any other enzyme, their activity is affected by a number of factors. Hence, in this study, the effect of enzyme and substrate concentrations on peroxidase activity of filtered extract of ground mango (Mangifera indica) kernel suspension (400 g L-1) was studied colorimetrically at 420 nm. Pyrogallol 5% w/v as substrate, filtered extract of crude enzyme from the kernel and phosphate buffer (pH 6.0) were employed in the analysis. Experimentally, first order reaction rate was observed with varied volumes of enzyme source up to 1.6 mL at a constant 1.0 mL of the substrate. On the other hand, holding the enzyme extract constant at 0.4 mL, peroxidase activity varied proportionately with substrate concentration up to 3.0 mL. Statistically, the mean of triplicate values were taken for each concentrations of enzyme and substrate taken. The result was subjected to regression analysis. From lineweaver-Burk plot, Km of 1.48 mM and Vmax of 0.29 units g-1 mango kernel were derived for peroxidase of mango (Mangifera indica) kernel using pyrogallol as substrate in this study. This study therefore identified limits of enzyme and substrate concentrations suitable to assay peroxidase activity. Hence, peroxidase in Mango (M. indica) kernel can be isolated, purified, characterised and quantitatively made available for industrial and medical use.