AbstractAs a new woody oleaginous crop, tree peony is now being widely developed and utilized, which attributed to the outstanding oil‐use features regarding the fatty acid profiles and bioactivity. The major fatty acid profiles and bioactivity of seed oils in ten tree peony varieties collected from a common garden were investigated in the present study. The results showed that the oil yields, fatty acid profiles, bioactivity of seed oils existed significant variations among ten tree peony varieties (P<0.05). Considered the application value, ‘Fengdan’ (FD) and ‘Ziban’ (ZB) was the optimal resources as the high oil yields, rich unsaturated fatty acids especially high α‐linolenic acid of 40.46 %, great antioxidant activity with low IC50 values, high ABTS and FRAP values, and strong antimicrobial activity with high DIZ and low MIC/MBC values. The study also confirmed seed oil of tree peony as the potential raw materials sources in functional food, pharmaceuticals and cosmetics for human health.