Recent observations suggest that products of nonparenchymal liver cells such as eicosanoids and cytokines might play a role in the expression of liver injury after administration of acetaminophen and other noxious agents. We therefore investigated the effect of a fish oil diet, which results in the generation of eicosanoids with altered biological properties and suppresses the production of certain cytokines on acetaminophen hepatotoxicity. Mice were fed a diet with either 20% fish oil containing n-3 fatty acids or 20% olive oil containing n-6 fatty acids for 2 wk. Cytochrome P-450 activity and the concentration of glutathione were similar in the two groups before acetaminophen administration. Nevertheless, 24 hr after the administration of 375 mg/kg acetaminophen intraperitoneally, the extent of centrilobular necrosis and the activity of ALT in plasma were significantly lower in the n-3 fatty acid group (median = 277 vs. 3,367 IUb, p < 0.001). In the n-3 fatty acid group covalent binding of the drug to liver proteins (0.19 2 0.03 vs. 0.67 f 0.07 nmol/mg protein; p < 0.01) and the median plasma concentration of acetaminophen (0.1 vs. 0.6 mmol/L) were significantly lower 3 hr after dosing. Mice fed the n-3 fatty acid diet excreted less acetaminophen sulfate but significantly more acetaminophen glucuronide in 24 hr. Thus the major protective effect of the fish oil diet appears to be an increased clearance of acetaminophen resulting from a stimulation of the glucuronidation of acetaminophen, which may be due to the fluidization of microsomal membranes by fish oil. (HEPATOLOGY 1991;13:557-561.) Liver injury caused by acetaminophen is due to the formation of a reactive toxic metabolite by the hepatic cytochrome P-450 system (1). A growing body of data suggests that the extent of hepatocellular necrosis not only depends on the amount of toxic metabolite formed and its detoxification by glutathione (2, 3) but also on secondary events occurring after arylation of macromolecules in hepatocytes. Chemotactic signals lead to an accumulation of neutrophils in the liver (4) and to hepatic congestion (5). Products of macrophages and