The aim of this work is to describe the role of different phospholipases in hepatic phospholipid catabolism. Therefore isolated rat hepatocytes enriched in labeled dilinoleoyl-, dipalmitoyl- or dimyristoylglycerophosphocholine were prepared by pulse incubation with [3H]glycerol and 14C-labeled fatty acid. The labeled cells were chased up to 4 h in a tracer-free medium and the degradation of different phosphatidylcholines studied. After a 2-h chase about 40% of dilinoleoyl-, 70% of dipalmitoyl- and 30% of dimyristoylglycerophosphocholine were degraded. From the positional distribution of 14C-labeled fatty acid and the change in the doubly labeled molecular species of phospholipids, it was concluded that tb degradation of dilinoleoylglycerophosphocholine and that of phosphatidylethanolamine could be accounted for by the action of phospholipase A1, while the degradation of dipalmitoylglycerophosphocholine proceeded through the action of phospholipase A2. Dimyristoylglycerophosphocholine was probably cleaved by the combined action of both phospholipases A1 and A2. Up to 10 mM tetracain, added to the chase medium, effectively blocked the action of both phospholipase activities. A considerable part of 2-linoleoyl- and 1-palmitoylglycerophosphocholine liberated during the chase was reutilized for phosphatidylcholine synthesis without further degradation.