The contribution of electrophilic and radicalic intermediates to phospholipid adducts formed by halomethanes in vivo

Abstract

The different production of phosgene and free-radicals from CHCl3 and CCl4 was determined in vitro and in vivo, by measuring the regioselective binding to the two intermediates to phospholipid (PL) molecules. Results clearly indicated that this assay can be successfully used to selectively detect electrophilic and radicalic metabolites produced in vivo and selectively quantitate their adducts. The in vivo biotransformation of CCl4, similarly to the in vitro situation, resulted in the formation of radicals only, the contribution of phosgene to the structural damage of PL being negligible. These findings allowed us to rule out the hypothesis of substantial formation of radicalic intermediates from CHCl3 in phenobarbital (PB)-pretreated Sprague-Dawley (SD) rats, derived from in vitro data. While the role of reduced glutathione (GSH) in preventing COCl2-derived damages seems to be less important in vivo than in vitro, it is not possible to rule out the action of radical scavenging systems in decreasing the level of adducts with fatty acyl chains (FC) of PL measured in vivo.

The role of different cytochrome P450 isoforms in in vitro chloroform metabolism

“…Possibly in contrast with these data, we have recently reported that the in vitro oxidative and reductive pathways of 5 mM CHCl 3 metabolism in Osborne Mendel rat liver microsomes were not affected by ethanol pretreatment of the animals, although the levels of CYP2E1 and its activity with model substrates were increased (18). Cytochrome P4502B1/2 (CYP2B1/2) also was shown to participate in chloroform metabolism since phenobarbital (PB) pretreatment enhanced the production of CO 2 in vivo (19) and of COCl 2 (15) in vitro, and it increased the in vivo covalent binding of chloroform metabolites to microsomal constituents (20,21). CYP2B1/2 was clearly involved not only in chloroform oxidation but also in chloroform reductive activation.…”

Characterization of a Phospholipid Adduct Formed in Sprague Dawley Rats by Chloroform Metabolism: NMR Studies

“…The binding of phosgene, the CHCl 3 oxidative metabolite, to the PL polar heads has also been shown in isolated microsomes and hepatocyte incubations and in vivo (6,7,29). In a previous in vitro study, we showed that this metabolite selectively binds to a specific PL in isolated hepatocytes (13).…”

Clinical Drug Treatment of Edemagenic Gas-Induced Lung Injury

“…Furthermore, phosgene has been shown to increase lipid peroxidation, cause the release of sulfidopeptide leukotrienes, alter important cell ultrastructures, and react directly with lung surfactant (22,24,27,43). In addition, it has been shown that phosgene can form adducts with renal microsomal phospholipid polar heads (19). In the lung, the end result of these disturbances is an increase in susceptibility to interstitial edema followed by alveolar flooding that can be potentially fatal for exposed individuals (23,44,79).…”

“…Phosgene also has been found to be toxic through normal metaboUc detoxification mechanisms unrelated to durect inhalation exposure. In hepatocytes, it was determined that the oxidative metaboUte of chloroform, phosgene, forms adducts with phospholipids such as phosphotidylcholine (PC) and phosphotidylethanolamine (PE) under hypoxic or normoxic conditions (19)(20)(21). Adduct formation could be mechanistically important during the injury process because alveolar surfactant is largely phospholipid in content and alveolar edema causes a locally hypoxic environment (22).…”