A comparative study has been made of the assimilation and metabolism of rac-1 and 2-[9, 10(-3)H]-octadec-9-enylglycerol in a clone of epithelial-like cells isolated from rabbit liver. Based on cell protein content, the free glycerol ether isomers attained equal cellular concentrations. As shown by isolation and degradation experiments, however, the incorporation of radioactive 1-monether was appreciably higher than that of radioactive 2-monoether in both the triacylglycerol and phospholipid fractions. The 1-monoether, unlike the 2-monether, was also a significant source of esterified fatty acids in both lipid fractions. In addition, the 1-monoether, but not the 2-monoether, was an active precursor of plasmalogens, particularly ethanolamine plasmalogen. In contrast to the 1-monoether, the 2-monoether was a more active precursor of triacylglycerols than it was of phospholipids. The results indicate that in the rabbit liver cells the pathway of complex lipid synthesis from 1-monoether was via 1-alkyl-sn-glycerol-3-phosphoric acid and from 2-monoether via 1-alkyl-2-acyl-sn-glycerol.