A quantitative method for the detection and validation of catalase activity at physiological concentration in human...

A novel method has been proposed to develop a simple, rapid, sensitive and affordable chromogenic attempt for the quantification of catalase (CAT) activity in blood samples. The method is based on the oxidation of pyrocatechol (PC) to give quinone form which by oxidative coupling with aminyl radical of 4-aminoantipyrine (4-AAP) resulting from H₂O₂/CAT to produce a pink colored quinone-imine product with λ_max = 530 nm in a 100 mmol/L of tris buffer of pH 9.8 at room temperature (30 °C). The linearity of CAT assay was between 0.316 and 10 U/mL. The accuracy ranges for CAT having concentrations of 1.25, 5 and 7.5 μmol/L were 89-105.52, 90-107%, and 91-104.58% respectively. Within-run and between-run precision studies showed CV's of 1.98-3.02% (n = 7) and 2.97-4.40% (n = 7), respectively. The detection and quantification limits of CAT were 0.12 and 0.225 μmol/L, respectively. The Michaelis-Menten constant and maximum velocity of the reaction was K_m = 1.052 mM and V_max = 0.168 μmol/min, respectively. The present method provides a convenient means for investigating the usefulness of CAT measurements in biological sample assessing the potential for free radical-induced pathology.