Bernhard H. Lauterburg

Determinants of hepatic glutathione turnover: toxicological significance

The tripeptide glutathione participates in a number of critically imporlant cellular processes (Fig. I). The function best known to the pharmacologist is associated with the nucleophilic structure of the compound which enables it to form conjugates with xenobiotics. The formation of these glutathione adducts proceeds spontaneously with some electmphilie compounds but in general conjugation is greatly facilitated by the glutathione transferases’. This group of enzymes binds a bmad spectrum of compounds and in this respect is very similar to serum albumin. In addition. however, the enzymes have a specific binding site for glutathione thereby bringing the nucleophilic glutathione in close proximity to an electmphilic atom of the first ~b strate. The resulting conjugates are 1” ‘li;cursors of the generic group of exck9 . products known as mercapturic acids. C.Ilowing removal of the gamm&glutamyl moiety by gamma-glutamyltransferase mainly located in the kidneys and intestinal mucosa. the glycine moiety is split off by peptidases and the resulting cysteine adduct of the original compound is then N-acetylated to the N-acetyl-cysteine thioether designated as mercapturic acid. The glutathione adducts are more polar than the original compc>unds, which facilitates their excretion. ln addition, when dealing with chemically reactive metatmlites of senobiotics such as acetaminophen. conjo gate formation prevents the covalent binding to tissue macromolecules and the hcpatotoxicity of the metabolitesY. The availability of glutathione for conjugate formation, therefore, is a critical determinant of the toxicity of electrophilic drug metabolites capable of alkylating nucleophilic sites on vital hepatic molecules.

Oxidation of hydrazine metabolites formed from isoniazid

Studies in rats indicate that the metabolic activation of acetylhydrazine, a metabolite of isoniazid, is a critical determinant of the hepatotoxicity of isoniazid. As demonstrated in that model, the formation of 14CO2 after the administration of 14C‐acetylisoniazid reflects the activity of the toxic pathway. A similar approach in man should make it possible to demonstrate the presence and to assess the quantitative importance of this toxifying pathway, and thus to evaluate its role in the pathogenesis of isoniazid hepatitis. We gave 300 mg isoniazid together with 10 µCi 14C‐acetylisoniazid (12 mg) to 17 healthy subjects and determined the time course of the plasma concentrations of isoniazid, acetylisoniazid, acetylhydrazine, and diacetylhydrazine and of the exhalation of 14CO2. The time course of 14CO2 in breath closely paralleled the plasma concentration‐time curve of acetylhydrazine but not those of acetylisoniazid or diacetylhydrazine, indicating that the 14CO2 originated directly from the metabolism of acetylhydrazine. The cumulative exhalation of 14CO2 increased with decreasing rate of acetylation of isoniazid, such that slow acetylators generated more 14CO2 than rapid acetylators. Simulation studies demonstrated that even if the data are corrected for the different formation of acetylisoniazid from isoniazid in slow and rapid acetylators, the slow acetylators still generated more 14CO2. The data therefore indicate that a substantial fraction of the acetylhydrazine formed from isoniazid passes through a pathway that has been shown in animals to generate highly reactive and hepatotoxic intermediates. The observation that slow acetylators metabolize more isoniazid via the toxic pathway, together with recent data showing an exceptionally high incidence of hepatitis in slow acetylators receiving large therapeutic doses of isoniazid, further support the hypothesis that the metabolic activation of acetylhydrazine from isoniazid is an important determinant of the hepatotoxicity of isoniazid in man.

Determination of isoniazid and its hydrazino metabolites, acetylisoniazid, acetylhydrazine, and diacetylhydrazine in human plasma by gas chromatography—mass spectrometry

Abstract A gas chromatographic—mass spectrometric assay for isoniazid and its hydrazino metabolites in human plasma was developed. The trimethylsilyl derivatives of diacetylhydrazine and acetylisoniazid and of the benzaldehyde hydrazones of acetylhydrazine and isoniazid were separated on a 1% OV-17 column and quantitated by single ion monitoring using a LKB 9000 mass spectrometer. Deuterated analogues served as internal standards. The method is well suited for the determination of the hepatotoxic hydrazino metabolites of isoniazid in human plasma following an oral therapeutic dose of isoniazid.

Alkylation and Peroxidation Injury from Chemically Reactive Metabolites

As initiators of chemical-induced tissue injury, alkylation and peroxidation reactions are often discussed as though they were mutually exclusive or competing adversaries for our love and affection, our hearts and souls. They are neither mutually exclusive nor competing hypotheses. In fact for some chemicals converted to radical metabolites, they might even be additive.

In Vivo Regulation of Hepatic Glutathione Synthesis: Effects of Food Deprivation or Glutathione Depletion by Electrophilic Compounds

The increased glutathione turnover that occurs during fasting results from two mechanisms. Because of a decrease in the intrahepatic free glutathione: mixed disulfide ratio, which is apparently mediated by c-AMP, the free glutathione pool contracts and turns over more rapidly in order to maintain glutathione synthesis. In addition, glutathione consumption via the gamma-glutamyl cycle apparently is increased during fasting. Additional data indicate that an inability to stimulate further the rate of glutathione synthesis during fasting, rather than a reduction in glutathione synthesis are previously postulated, probably explains the increased susceptibility of fasted animals to tissue attack by electrophilic drug metabolites.

Drug-induced liver injury.

In the usual course of events, the liver detoxifies hepatocytotoxic intermediates of drug metabolism. But when protective systems are overloaded by normally harmless drugs, the intermediates can cause massive, even fatal, hepatic necrosis.