The porphyrinogenic agent 3,5-dicarbethoxy-2,6-dimethyl-4-ethyl-1,4- dihydropyridine (DDEP) is known to destroy hepatic cytochrome P450 isoforms in many animal species by ethylating the heme moiety by mechanism-based inactivation process. DDEP appears to have singled out three P450 isoforms (P450 2C11, 2C6 and 3A) for this inactivation. DDEP- mediated inactivation of P450 2C11 and 2C6 appears to entail destruction by N-ethylation to the prosthetic heme. On the other hand, that of P450 3A appears to involve destruction of the prosthetic heme to a reactive species that covalently bounds to the apoprotein. However, the exact mechanism of DDEP-mediated destruction of the prosthetic heme of P450 3A isoform is virtually unknown. Intramolecular isotope effects associated with oxidation of 3,5- dicarbethoxy-2,6-dimethyl-4-[4-d,4-H]dihydropyridine (DDDP) catalyzed by P450 2C11 and 3A4 were 1.46 and 1.98, respectively. These results indicate that suicide inactivation of both P450 isoforms is initiated by one electron oxidation of nitrogen of DDEP. Horseradish peroxidase (HRP) is used as a model system for DDEP-mediated inactivation of P450 3A. Two major metabolites of DDEP catalyzed by HRP in the presence of either hydrogen peroxide or iodosobenzene are 3,5- dicarbethoxy-2,6-dimethyl-4-ethyl-pyridine (EDP) and 3,5-dicarbethoxy- 2,6-dimethyl-pyridine (DP). Product ratios of these metabolites changes in a pH dependent manner, and the pathway leading to the formation of DP is solely responsible for the inactivation of HRP. An altered heme product appeared corresponding to the loss of prosthetic heme of HRP. Plasma desorption mass spectrometry (PLMS) revealed that this altered heme (mass 663) is formed by addition of an ethyl group and a hydroxy group to heme.
