The hydrogen peroxide-oxidation of o-phenylenediamine (OPD) catalyzed by horseradish peroxidase (HRP) at 37 degrees C in 50 mM phosphate buffer (pH 7.0) was studied by calorimetry. The apparent molar reaction enthalpy with respect to OPD and hydrogen peroxide were -447 +/- 8 kJ mol(-1) and -298 +/- 9 kJ mol(-1), respectively. Oxidation of OPD by H2O2 catalyzed by HRP (1.25 nM) at pH 7.0 and 37 degrees C follows a ping-pong mechanism. The maximum rate V-max (0.91 +/- 0.05 mu M s(-1)), Michaelis constant for OPD K-m.S (51 +/- 3 mu M), Michaelis constant for hydrogen peroxide K-m,K-H2O2 (136 +/- 8 mu M), the catalytic constant k(cal) (364 +/- 18 s(-1)) and the second-order rate constants k(+1) = (2.7 +/- 0.3) x 10(6) M-1 s(-1) and k(+5) = (7.1 +/- 0.8) x 10(6) M-1 s(-1) were obtained by the initial rate method. (c) 2006 Elsevier B.V. All rights reserved.