It has been previously shown that the positional isomers of triacylglycerol (TAG) containing palmitic acid (P) and highly unsaturated fatty acids (HUFAs) such as DHA (D) and EPA (E) vary between fishes and marine mammals. However, it has not yet been understood why in marine mammals HUFAs are located only at the α position when two palmitic acid chains combine, and not in fishes. In order to gain further understanding of the biosynthetic pathways involved in the formation of these asymmetric TAGs, we investigated whether the HUFA in the TAG of marine mammals exists predominantly at the sn-1 or sn-3 position. We examined the TAG positional isomers and enantiomers in marine organisms in detail. As a result, while PDP and PEP were not detected, sn-PPD and sn-PPE were found in abundance in marine mammals. For fishes, on the other hand, PDP, PEP, sn-PPD, and sn-PPE were all identified. In the case of TAGs that contain two HUFAs and one palmitic acid, marine mammals were rich in DPD and EPE whereas fishes were rich in sn-PDD and sn-PEE.