We reported that phospholipids of interleukin-2 (IL-2)-dependent murine cytotoxic T lymphocytes (CTL) contained high amounts of ether lipids (Biochem. Mol. Biol. Int., 33, 137-146, 1994). In this study, the molecular species of fatty acids at the sn-1 and sn-2 positions were determined. First, phosphatidylcholine mainly consisted of the 1,2-diacyl subclass (78%) in which the major molecular species was 16:0/18:1 (24%). It should be noted that arachidonic acid was abundant in both 1-O-alkyl-2-acyl subclasses in the form of 18:0/20:4 (21%) even though the content of the subclass was low (8%). Second, phosphatidylethanolamine mainly consisted of ether subclasses (93%) and contained a large amount of arachidonic acid in the forms of either 18:0/20:4 (19%) or 16:0/20:4 (10%). Third, phosphatidylinositol, a putative phospholipid involved in transmembrane signal transduction, had a significant amount of arachidonic acid at the sn-2 positions of 1,2-diacyl and 1-O-alkyl-2-acyl subclasses. Fourth, phosphatidylserine, consisting of mostly the 1-O-alkyl-2-acyl subclass (86%), contained arachidonic acid dominantly in the form of 18:0/20:4 (33%). Considering these results, it appears that CTL have the potential to mediate its signal transduction mediated by the bioactive lipids, i.e. arachidonic acid metabolites and the ether lipids.