Biological Oxidation of Nitrogen in Organic Compounds and Disposition of N-Oxidized Product

Abstract

N-Oxidation of nitrogenous compounds has been shown to be of special biological interest: many N-oxidized aromatic amines have been recognized to be ultimate mutagens and carcinogens; ferrihemoglobin formation is mediated by a series of N-oxidized products, and allergic disorders arise from interaction of such compounds with cellular macromolecules. There are several types of nitrogen containing compounds which undergo N-oxidation in biological systems. These include aliphatic and aromatic primary, secondary, and tertiary amines, heterocyclic secondary amines, heteroaromatic and alicyclic tertiary amines, primary and secondary amides as well as imines, hydrazines, and azo compounds. N-Oxidation products tend to be formed in small amounts and are often labile. Colorimetric methods of analysis, although frequently used, tend to be nonspecific and other products may interfere with the measurement. Chromatographic methods allow direct identification and quantification and take into account the lability of these metabolites. Direct quantification of N-oxidation products by GLC has been achieved by derivatisation to give more stable compounds. Recently HPLC has been successfully employed to resolve some of the analytical problems. The natural occurrence of a large variety of N-oxidized compounds in plant and animal tissues has been recognized. Similarly, a broad spectrum of nitrogenous drugs and other xenobiotics are converted to the corresponding N-oxidation products after in vivo administration to men and animals. These include analgesics, narcotics, psychotherapeutic drugs, sympathomimetics, chemotherapeutic agents, and carcinogens. The extent of N-oxidation of the individual amine compounds has been found to vary with the species investigated. Organ specificity and genetic control are further factors to be considered. The site of N-oxidation of amine compounds has been detected to be located in the endoplasmic reticulum of the cell. By using isolated hepatic microsomal fractions, in vitro experiments demonstrated the N-oxidation process to exhibit typical mixed-function oxidation characteristics. Nevertheless, there is still considerable controversy as to the enzymology of N-oxidation. Experiments with classic inhibitors and modifiers of the cytochrome P-450 mixed-function oxidase system suggest the N-hydroxylation of primary alkyl- and arylamines to be a hemoprotein-dependent reaction. N-Oxidation of secondary and tertiary alkyl- and arylamines is believed to proceed by a cytochrome P-450-independent pathway represented by a FAD-containing flavoprotein oxidase. However, evidence has accumulated demonstrating some involvement of cytochrome P-450 in the N-oxidation of N,N-dialkylanilines, pyridines, and some alicyclic tertiary amines. The hemoprotein system also appears to mediate N-oxidation of acetamides, imines, and monoalkylhydrazines. Steric features of the individual amine compounds have been found to influence the N-oxygenating capacity of the enzyme systems. Basicity and nucleophilicity of the amine substrates has been taken to predict whether the preferred route of N-oxidation is via the cytochrome P-450 system or the flavoprotein amine oxidase. The value of such concept is discussed. Other mechanisms such as the peroxidatic transformation of aniline to phenylhydroxylamine by catalase and chloroperoxidase are discussed. N-Oxidation products undergo further metabolism by various enzyme systems. The most important reactions are: (1) Reduction of hydroxylamines, nitro compounds, and N-oxides; (2) oxidation of hydroxylamines and oxims; (3) oxim-amide rearrangement; (4) N-oxide dealkylation; and (5) N-hydroxyacetamide glucuronidation, sulfation, acylation, and deacetylation. The latter transformation reactions effect special reactivity of the metabolites with tissue macromolecules.