New Insight into the Mechanism of NADH Model Oxidation by Metal Ions in Nonalkaline Media.

ABSTRACT

For a long time, it has been controversial that the three-step (e-H+-e) or two-step (e-H•) mechanism was used for the oxidation of nicotinamide adenine dinucleotide coenzyme (NADH) and its models by metal ions in nonalkaline media. The latter mechanism has been accepted by the majority of researchers. In this work, 1-benzyl-1,4-dihydronicotinamide (BNAH) and 1-phenyl-l,4-dihydronicotinamide are used as NADH models and ferrocenium (Fc+) metal ion as an electron acceptor. The kinetics for oxidation of the NADH models by Fc+ in pure acetonitrile was monitored by using UV-vis absorption and a quadratic relationship between kobs and the concentrations of NADH models was found for the first time. The rate expression of the reactions developed according to the three-step mechanism is quite consistent with the quadratic curves. The rate constants, thermodynamic driving forces, and kinetic isotope effects of each elementary step for the reactions were estimated. All results supported the three-step mechanism. The intrinsic kinetic barriers of the proton transfer from BNAH+• to BNAH and the hydrogen-atom transfer from BNAH+• to BNAH+• were estimated by using Zhu equation; the results showed that the former is 11.8 kcal/mol and the latter is larger than 24.3 kcal/mol. It is the large intrinsic kinetic barrier of the hydrogen-atom transfer that makes the reactions choose the three-step rather than two-step mechanism. Further investigation of the factors affecting the intrinsic kinetic barrier of chemical reactions indicated that the large intrinsic kinetic barrier of the hydrogen-atom transfer originated from the repulsion of positive charges between BNAH+• and BNAH+•. The greatest contribution of this work is the discovery of the quadratic dependence of kobs on the concentrations of the NADH models, which is inconsistent with the conventional viewpoint of the "two-step mechanism" on the oxidation of NADH and its models by metal ions in the nonalkaline media.