Azo dyes are of concern due to their harmful effects on the environment and human health. The oxidation of 2,2'-dihydroxyazobenzene (DHAB) catalyzed with recombinant Coprinus cinereus (rCiP) peroxidase was investigated. The kinetic measurements were performed using the spectrophotometric and fluorimetric methods. The dependences of the initial reaction rates on enzyme, substrate and hydrogen peroxide concentrations during DHAB oxidation were established, and bimolecular constants of enzyme interaction with DHAB were calculated. This research demonstrated that the initial biocatalytic oxidation rates of DHAB depend on the pH and the estimated pKa values of the active forms of rCip. This study's findings thus contribute to a more comprehensive understanding of the biocatalytic oxidation of DHAB, providing valuable data for assessing the long-term toxicity, carcinogenesis and epigenetic effects of azo dyes in the environment.