Oxidation of substituted N,N-dimethylanilines by cytochrome P-450: estimation of the effective oxidation-reduction potential of cytochrome P-450

Abstract

Rates of N-demethylation of N,N-dimethylaniline and of eight meta- or para-substituted N,N-dimethylanilines by rat liver cytochrome P-450PB-B (P-450) were determined under conditions where oxidation was supported by iodosylbenzene or NADPH-P-450 reductase. The rates of dimethylaniline oxidation were found to correlate with the substrate oxidation-reduction potential within each series of substrates supported by a particular oxygen activation protocol; the kcat for each substrate studied was approximately 20-fold faster in the iodosylbenzene-supported system relative to the NADPH-P-450 reductase supported system. Since the N-demethylation of amines is believed to proceed via an initial electron-transfer step, a kinetic scheme for P-450 was proposed that enabled evaluation of the data according to theoretical treatments that correlate rates of electron transfer with extrakinetic parameters. In these analyses, the data could be fitted to the Rehm-Weller and Agmon-Levine equations, providing lambda values (for the energetics of enzyme-substrate reorganization) of 22-26 kcal mol-1 and apparent E1/2 (oxidation-reduction potentials) of 1.7-2.0 V (vs saturated calomel) for the oxidized enzyme. The apparent E1/2 for the enzyme is composed of contributions from the intrinsic potential of the active prosthetic core of the enzyme, the Fe = O - porphyrin species, and a coulombic factor that is a function of the charge-separated radical anion/radical cation pair produced upon electron transfer.(ABSTRACT TRUNCATED AT 250 WORDS)

Mechanism of 17α,20-Lyase and New Hydroxylation Reactions of Human Cytochrome P450 17A1

“…H 2 O 2 (to 10 mM) or iodosylbenzene (to 2 mM) was added at varying concentrations (from aqueous stocks). The incubations were done for 5 min with H 2 O 2 and for 30 s with iodosylbenzene (41,92), with extraction into CH 2 Cl 2 and analysis of the conversion of 17␣-hydroxyprogesterone to androstenedione and of 17␣-hydroxypregnenolone to DHEA by UPLC. The ⌬ 5 steroids were converted to ⌬ 4 steroids by treatment with cholesterol oxidase prior to LC-UV analysis (37).…”

Recent density functional theory model calculations of drug metabolism by cytochrome P450

“…The N-dealkylation reaction of amine has been the subject of several experimental and theoretical investigations [85][86][87][88][89][90][91][92][93][94][95][96][97][98][99][100][101]. NDealkylation of amine is proposed to result from hydroxylation of a carbon adjacent to the N atom, forming the unstable carbinolamine, which decomposes subsequently to give rise to a carbonyl product and a dealkylated amine.…”

“…One is a hydrogen atom transfer (HAT) reaction [86][87][88][89], labeled as (i), on the carbon adjacent to the N atom, followed by oxygen rebound to form the carbinolamine, which decomposes subsequently to formaldehyde and a dealkylated amine. An alternative mechanism involves a single electron transfer (SET) on the N atom itself [90,91,98], labeled as (ii), followed by proton transfer, and then oxygen rebound to form the carbinolamine, which then decomposes as outlined above.…”

“…Experimental results [87][88][89]99] on N-cyclopropylamines fit the HAT mechanism as well. The SET mechanism is supported by the low KIEs of oxidative dealkylation [93,94,101], as well as the correlation of the reaction rates for N-demethylation of p-X-DMAs with the Hammett substituent constant para and the redox potential E 1/2 of the p-X-DMA substrates [90,91].…”

Mechanisms of cytochrome P450 substrate oxidation: MiniReview

“…When substrates have atoms with low-oxidation potentials (e.g., N, S, P) within an accessible distance, the FeO 3+ can act as an oxidative electrode; the oxidation potential can be estimated to be >1 V [13]. With alkyl-substituted heteroatoms, the FeO 2+ species appears to act as a base to help remove an α-proton, and oxygen rebound to the incipient carbon radical yields a carbinolamine or the equivalent oxidized heteroatom species [14,15].…”