Two groups of rats (200 g, male) were fed, respectively, a choline-deficient and a choline-supplemented diet for 3 days. On the 3rd day each rat was injected intravenously with rat serum containing 9,10-3H-palmitate and 18-14C-stearate. The fate of the labelled fatty acids was followed in the triglycerides and phospholipids of liver and serum for time periods ranging from 1 min to 6 h. When compared to the choline-supplemented controls, the livers in the choline-deficient rats were fatty and were characterized by: (1) an increased hepatic triglyceride pool, (2) a decrease in turnover of hepatic triglycerides with sequestration of fatty acids and decrease in their recycling, (3) an increase in the hepatic free fatty acid pool and total phospholipid pool, the latter reflecting the net result of synthesis and transport, (4) a decrease in the early and rapid turnover of the phosphatidylcholine pool, (5) a decrease in turnover of the phosphatidylethanolamine pool, (6) a decrease in transport of triglycerides and phospholipids from liver to serum, and consequently, (7) a decrease in triglyceride and phospholipid pools in serum. These results confirm previous observations that, although an impairment of transport of both triglycerides and phospholipids from liver to serum exists in choline-deficient rats, the hepatic accumulation of triglycerides is far greater than that of phospholipids. This effect is due to the presence of a potential triglyceride pool in liver that can accommodate large amounts of triglyceride within a short period of time.