Abstract 2-Ethylhexanol, a metabolite of the commonly used plasticizer di(ethylhexyl)phthalate, was shown to cause toxicity exclusively to periportal regions of the perfused liver (Keller et al., 1990, J. Pharmacol. Exp. Ther., 252, 1355–1360.) To determine whether this toxicity was due to local oxygen tension or to drug delivery, isolated cylinders (plugs) of periportal and pericentral regions of the liver lobule from rats pretreated with phenobarbital were collected with a micropunch following brief perfusion of the organ. Plugs were 0.2 mm wide and 0.5 mm long and weighed between 0.5 and 1 mg each. Following incubation for at least 2 hr in Eagle's medium, they were judged viable based on production of urea at high rates and minimal leakage of lactate dehydrogenase (LDH). Plugs could be cultured for up to 24 hr with minimal loss of activity. Urea synthesis from ammonium chloride (3 m m) by plugs incubated in Krebs-Henseleit buffer equilibrated with 95% O 2:5% CO 2 was proportional to protein concentration and was linear with time for up to one hour at rates around 75 μmol/g/hr. Icubation of plugs with 2-ethylhexanol (0.1 to 3 m m) diminished urea synthesis in a dose-related manner (half-maximal effect = 0.5 m m). Ethylhexanol also caused extensive cell damage assessed from LDH leakage in incubations at 800 μ m O 2 but significantly less injury at 200 μ m O 2. Concomitantly, urea synthesis was inhibited by ethylhexanol by over 80% at 800 μ m O 2 but less than 50% at 200 μ m O 2. Plugs isolated from both regions of the liver lobule were affected similarly by ethylhexanol and O 2. Taken together, these data indicate that ethylhexanol toxicity is dependent on oxygen tension in isolated sublobular regions of the liver lobule, and therefore it is unlikely that drug delivery can explain the selective injury to periportal regions in studies with the perfused liver.