AN EVALUATION OF THE ROLES OF METABOLIC DENITROSATION AND ALPHA-HYDROXYLATION IN THE HEPATOTOXICITY OF N-NITROSODIMETHYLAMINE

N-Nitrosodimethylamine (NDMA) is a potent hepatotoxic agent in rats. N DMA causes cell death that does not correlate with known mechanisms of toxicity such as the production of oxidative stress or covalent bindi ng to proteins. The following studies were designed to determine wheth er NDMA cytotoxicity is the result of metabolic denitrosation or alpha -hydroxylation of the nitrosamine. We determined the toxicity of vario us metabolites of NDMA in monolayer cultures of primary rat hepatocyte s. NDMA was toxic at 0.1 mM in our cultures, but the metabolites forma ldehyde, methanol, and methylamine were not toxic at this concentratio n. We used diazeniumdiolates that spontaneously release nitric oxide ( NO) in aqueous media to deliver NO to hepatocytes in culture. The resu lts show that, while NO released from diazeniumdiolates causes death i n hepatocytes, the levels of NO produced during NDMA metabolism are in sufficient to account for the toxicity of the nitrosamine. NDMA-dB, th e fully deuteriated form of NDMA that undergoes approximately twice as much denitrosation in vivo as NDMA, was significantly less cytotoxic than NDMA. In contrast, N-nitroso-(acetoxymethyl)methylamine (AcO-NDMA ), a stable precursor of the methanediazonium ion, was found to cause toxicity equivalent to NDMA on a molar basis. Altogether, our results with methylamine, formaldehyde, methanol, the diazeniumdiolates, and N DMA-d(6) indicate that NDMA toxicity is not the result of metabolic de nitrosation, while the toxicity of AcO-NDMA provides strong evidence t hat the formation of the methanediazonium ion via alpha-hydroxylation leads to cell death.