Binding of metabolically derived acetaldehyde to hepatic proteins in vitro.

Homogenates of rat liver incubated with 10 mM [14C]ethanol were analyzed for acetaldehyde production and for both stable and unstable radiolabeled acetaldehyde adducts with proteins. During incubation, formation of acetaldehyde and of 14C-labeled proteins both increased with time in a parallel manner. Acetaldehyde generation and subsequent formation of radiolabeled proteins were potentiated by supplementation of the cell-free system with NAD+ (1 mM). Cycloheximide (0.1 mM) caused no significant reduction in protein-bound radioactivity, whereas the addition of strong nucleophiles L-cysteine (5 mM) and reduced glutathione (5 mM) each decreased radiolabeling by 50 to 60%. Preheating of crude homogenates at 90 degrees C, prior to incubation with [14C]ethanol profoundly decreased subsequent production of acetaldehyde and formation of 14C-labeled proteins. The results indicate that the major source of protein-bound radioactivity derived from [14C]ethanol oxidation in this system is due to binding of enzymatically derived [14C]acetaldehyde to hepatic proteins.