Cytochrome P-455 nm complex formation in the metabolism of phenylalkylamines. VII. Enzyme interactions with synthetic 2-nitroso-1-phenylalkanes.

Cytochrome P-455 nm complex formation in rat liver microsomes was investigated with 2-nitroso-1-phenylpropane, the nitroso compound related to amphetamine, and six homologues comprising different alpha-alkyl substituents. The C-nitroso compounds were synthesized and obtained as trans nitroso dimers, the only form in which they are available in pure solid state. Their physical and chemical properties were investigated and their decomposition in ethanol solutions was correlated with the complexing efficacies of these solutions. Dissociation of the nitroso dimers to monomers constitutes an equilibrium which is displaced in favour of the dimer but with the monomers ultimately undergoing tautomerization to oximes. Based on these kinetics a mathematical model was produced, which by computer simulations gave simultaneously the dimer (Mo2), monomer (Mo) and oxime (Ox) concentrations of the substrate solutions used in the complex formation studies. The results show that the formation of cytochrome P-455 nm complexes is directly related to the numerically predicted concentrations of the nitroso monomers, but is very slow or absent when the dimer or oxime concentrations are at their highest. Substrate solutions derived from nitroso dimers with larger alpha-alkyl substituents (3-4 carbons) were devoid of complexing activities because of low solubility and very slow chemical kinetics.

Cytochrome P-455-nm complex formation in the metabolism of phenylalkylamines. VI. Structure--activity relationships in metabolic intermediary complex formation with a series of alpha-substituted 2-phenylethylamines and corresponding N-hydroxylamines.

The formation of cytochrome P-450 metabolic intermediary (MI) complexes from a homologous series of alpha-substituted 2-phenylethylamines and corresponding N-hydroxylamines was investigated during NADPH-dependent metabolism in liver microsomes from phenobarbital-pretreated rats. The alpha-alkyl substituent consisted of branched and unbranched alkyl chains ranging from 0-4 carbons and the benzyl group. All the compounds but 2-phenylethylamine generated the complex and double-reciprocal plots of the highest observed rate of complex formation vs. substrate concentration gave linear relations over a defined substrate range. The Vmax(obs) values for complex formation by the amines increased markedly with increasing size of the alkyl group and a good correlation was obtained between log Vmax(obs) and the logarithm of the octanol/buffer partition coefficient of the substrates. With the N-hydroxy compounds, complex formation was a much less selective phenomenon and without exception the rates were quite high with Vmax(obs) values as much as 100 times greater than those of the amines. The disappearance of substrate amines was independent of structure and at an initial substrate concentration of 100 microM about a 50% decrease was noted during a 20-min period in all cases. The results substantiate the previous notion that N-oxidation is a prerequisite for MI-complex formation from primary amines. The results also suggest that C- and N-oxidation have different rate-limiting steps and the microsomal enzymes catalyzing the N-oxidation seem to be deeply submerged in the lipid matrix, as amines with a low distribution are inactive or poor substrates for generating the cytochrome P-450 ligand. Also, it is evident that the MI complex formation does not impair the overall metabolism of the amines.

Microsomal formation and chemical decomposition of pargyline N-oxide.

Pargyline undergoes metabolic N-oxidation in rat and rabbit liver microsomal preparations. The reaction requires oxygen and is NADPH dependent. N-oxidation and N-demethylation are equal in both control and induced rat liver microsomes, while N-oxidation is more dominant in rabbit tissue. Experiments investigating the CO-sensitivity and the effects of metyrapone suggest that cytochrome P-450 systems are involved in both reactions in the rat while an additional enzyme is responsible for the N-oxidation in the rabbit. Pargyline N-oxide is characterized by chemical instability and undergoes two consecutive rearrangements to yield propenal and Schiff bases, the latter undergoing hydrolysis to aldehydes and primary amines. Accordingly, due to the inherent instability of the N-oxide, metabolic N-oxidation of pargyline is, in addition to alpha-carbon oxidation, indicated as a metabolic route to benzaldehyde. Similarly the ease with which pargyline N-oxide generates propenal implicates N-oxidation as a metabolic route to be considered when evaluating the toxicity of pargyline.