Xenobiotic metabolism in humans during early pregnancy: peroxidase-mediated oxidation and bioactivation of 2-aminofluorene

Abstract

Arylamines such as 2-aminofluorene (2-AF) are known teratogens and transplacental carcinogens in laboratory animal species. Although exposure of women to arylamines is likely to occur during pregnancy, how these chemicals are metabolized by the enzymes from the human conceptual tissues is currently unknown. Highly purified preparations of peroxidase isolated from human intrauterine conceptual tissues at 8 weeks of gestation were used to study in vitro metabolism of 2-AF. The oxidation of 2-AF was examined spectrophotometrically whereas the bioactivation was assessed from the covalent binding to protein and DNA using [3H] 2-AF. Using guaiacol as a model substrate, the purified preparations of peroxidase used exhibited a specific activity of 15-20 micromol/min/mg protein. 2-AF oxidation was found to be enzymatic in nature. Kinetic data obtained under optimal assay conditions yielded a Km = 41 microM for 2-AF, 8.33 microM for H2O2, and a Vmax=1.2 micromol 2-AF oxidized/min/mg protein. Under optimal assay conditions, the covalent binding of reactive intermediates to protein and DNA (nmol equivalent/min/mg enzyme/mg bovine serum albumin or calf thymus DNA) was observed at the rate of about 3.75 +/- 0.39 and 1.90 +/- 0.11 respectively. A significant decline in the rate of both oxidation and bioactivation of 2-AF was observed in the presence of classical peroxidase inhibitors, KCN and NaN3.

Role of Biotransformation in Conceptal Toxicity of Drugs and Other Chemicals

“…The importance of these findings is heightened by the observations that the same or similar enzyme capable of xenobiotic oxidation also occurs in human intrauterine tissues during the organogenesis ( 4-6 weeks of gestation) [213] and early fetal (10 weeks of gestation) [98] periods. Quantitatively, it is noteworthy that the partially 10 Wk Fetal [98] 3,3'-Dimethoxybenzidine Term placenta [209] 4-6 Wk Conceptus [213] Pyrogallol Term placenta [209] 4-6 Wk Conceptus [213] Tetramethyl phenylenediamine Term placenta [209] 4-6 Wk Conceptus [213] 10 Wk Fetal [98] Tetramethylbenzidine Term placenta [209] 4-6 Wk Conceptus [213] ABTS** Term placenta [209] 4-6 Wk Conceptus [213] Benzidine Term placenta [209] 4-6 Wk Conceptus [213] 10 Wk Fetal [98] Epinephrine Term placenta [209] 4-6 Wk Conceptus [213] Bilirubin Term placenta [209] 4-6 Wk Conceptus [213] p-Phenylenediamne Term placenta [209] 4-6 Wk Conceptus [213] 2-Aminofluorene Term placenta [208] 4-6 Wk Conceptus [213] 8 Wk Conceptus [214] purified conceptal peroxidase can catalyze oxidation of model substrates with several fold greater efficiency than the term placental enzyme [98,213]. Purified human fetal tissue peroxidase can also bioactivate the teratogen, 2-AF [214].…”