Using a liver ischemia model, we examined the change that occurs in the phospholipids and marker enzyme activities of subcellular membrane-structures, as well as in the ultrastructure of the cells during ischemia. The left and median lobes of liver were made ischemic by occlusion of branches of the hepatic artery and portal vein; after 3 h the liver was excised, homogenized and fractionated. Of total 79 rats, 29 rats received via gastric tube 100 mg chloroquine/kg for 7 days prior to ischemia, or an intraperitoneal injection of Nifedipine (50 μg/kg; n=23). In some of these rats (n=28), the occlusion was removed, and the analyses were carried out 24 h later. The following findings were obtained; 1) Ischemia of 3-h duration reduced the phospholipid content (per mg protein) of the lysosomal, microsomal and postnuclear fractions, but not that of the mitochondrial fraction. 2) Pretreatment with chloroquine prevented to a large extent the ischemia-induced reduction of phospholipids. Some phospholipids in the heavier-density lysosomes decreased, and some of those in the lighter-density lysosomes increased by ischemia, suggesting that ischemia changed the density distribution of lysosomal population. 3) Nifedipine also reduced the ischemia-induced phospholipid change but to a lesser extent. 4) Electron microscopic observation indicated that the number of lamellar bodies appeared in the chloroquine-treated rat liver was decreased during 3-h ischemia; the ultrastructural alterations of ischemia appeared not to be greatly influenced by the chloroquine pretreatment. 5) The activity of mitochondrial cytochrome c oxidase and that of lysosomal acid phosphatase were decreased by ischemia; the chloroquine treatment prevented these decreases, but Nifedipine treatment did not. The depressed activity of microsomal NADPH cytochrome c reductase was ameliorated by chloroquine. These results indicate that enzyme activities change independently of a change in phospholipid content in the subcellular organelles during ischemia. 6) The three enzyme activities remained depressed in the liver of the non-treated rat 24 h after the resumption of blood flow. At this time the mitochondrial phospholipid content was significantly decreased. These changes were largely prevented by pretreatment with cloroquine. 7) These results suggest that the Ca2+-activation of subcellular phospholipases or the action of lysosomal hydrolases cannot be the sole mechanism leading to the development of ischemic cell injury.