Structured lipid (SL) was synthesized from pine nut oil (PNO) and palm stearin (PS). In SL, 8.81% of Δ5-unsaturated polymethylene interrupted fatty acids (Δ5-UPIFAs) were intentionally incorporated into the sn-2 position through acyl migration. The obtained SL contained mostly the β′ form and a wide plastic range with solid fat index of 26.5% at 10°C to 2.29% at 35°C, indicating that the obtained SL (containing zero-trans fatty acid) may be desirable for soft (tub) margarine fat. Subsequently, the antioxidative effects of α-tocopherol (α-TOH), ascorbyl palmitate (AP), quercetin (Qu), and combinations thereof on SL were investigated. Results showed that Qu (500μg/g) showed the most effective antioxidant activity, followed by AP (500μg/g); while α-TOH with any concentrations (100, 200 and 500μg/g) did not show significant protective activity in the obtained SL. Each mixture of AP+Qu and α-TOH+AP+Qu also showed effective antioxidant activity in the obtained SL.