Spectral evidence for 2,2,3-trichloro-oxirane formation during microsomal trichloroethylene oxidation.

During aerobic incubation of trichloroethylene with rabbit liver microsomes and NADPH a difference absorption peak appears at 451-452 nm. Trichloroethylene does not form a ligand absorption spectrum with hepatic microsomes reduced by dithionite, or in anaerobic incubates in the presence of NADPH. Addition of trichloroethylene epoxide (2,2,3-trichloro-oxirane) to reduced suspensions of rabbit liver microsomes produces high difference absorption at 452 nm, the optical Ks being approximately 2 mM. Of all possible metabolites of trichloroethylene only trichloroethanol forms absorption in the vicinity of 480 nm, and the broad absorption band reveals relatively low absorption near 450 nm. Dichloroacetyl chloride is the main thermal rearrangement product of trichloroethylene epoxide, and also produces 452 nm absorption in reduced microsomes. However, the difference absorption is 5 times smaller than the absorption produced by the intermediate formed during incubation of trichloroethylene in metabolising liver microsomes. These observations include strong evidence for epoxide formation during microsomal oxidation of trichloroethylene. 14C-labelled trichloroethylene binds irreversibly to hepatic macromolecules in vivo and in vitro. Possible rearrangement pathways of 2,2,3-trichloro-oxirane and reactive intermediates are presented.

Irreversible binding of 3-14C-antipyrine to hepatic protein in vivo and in metabolizing liver microsomes.

After i.p. injection of 3-14C-antipyrine (10 micronmole = 1.9 mg with 10 micronCi per 10 g of body weight) to mice radioactivity was irreversibly bound to liver proteins. The irreversible binding reached maximal values of 0.15 nmole/mg protein in liver microsomes after 30-60 min. During 60 min incubation with liver microsomes of mice and rabbits (phenobarbital pretreated) and a NAKPH-regenerating system 3-14C-antipyrine was irreversibly bound to microsomal protein at a rate of 1.5 nmole/mg protein (mouse) and 3 nmole/mg protein (rabbit). In identical incubates with rabbit liver microsomes the 4-hydroxylation of antipyrine was 24 nmole/mg protein in 60 min and formaldehyde production from antipyrine 3 nmole/mg protein in 60 min. In incubates with rabbit liver microsomes the binding rate was 80-90% inhibited by 1mM metyrapone, SKF 525-A and trichloropropene epoxide respectively; 4-hydroxylation was 70-80% inhibited by the same substances. In the presence of 1mM GSH, cysteine or ethylene diamine binding was 30-40% inhibited, whereas 4-hydroxylation showed no inhibition.