Abstract: Using pumpkin seed oil (PSO) as the core material, pumpkin seed protein nanoparticles (PSP) and modified flaxseed gum (MFG) as the wall materials of microcapsules, pumpkin seed oil microcapsules were prepared by freeze-drying method through different combinations and proportions. The effects of different PSP/MFG mass ratios on the interfacial tension, storage stability, stability, encapsulation efficiency, microstructure, and in vitro digestion release characteristics of pumpkin seed oil microcapsules were investigated. The results showed that compared with the control group (P4), the stability of pumpkin seed oil microcapsules with different PSP/MFG mass ratios was significantly changed. Among them, when the PSP/MFG mass ratio was 4:5 (P4F5), the microcapsule size was the smallest (199.06 μm), the interfacial tension was the lowest (22.03 mN/m), and the three-phase contact angle was 89.665°, showing the best surface wettability and stability. PSP/MFG wall materials could effectively encapsulate PSO, with an encapsulation efficiency of 96.5%, and the highest encapsulation efficiency after storage for 30 days was 88.8%. The TSI value was 0.35, indicating the best stability. The in vitro digestion release characteristics of PSP/FG microcapsules showed that the stable pumpkin seed oil microcapsules had the function of controlling the release of pumpkin seed oil, which was characterized by a lower release rate in simulated gastric juice and a significant increase in release rate in simulated intestinal juice, thus effectively releasing pumpkin seed oil. By evaluating the performance of the microcapsule wall materials, it was indicated that the pumpkin seed protein-modified flaxseed gum composite wall material could be used as a good nano-level stable delivery carrier for pumpkin seed oil and improve its bioavailability. At the same time, it provided a theoretical reference for the deep processing of pumpkin seed oil and its comprehensive utilization in the fields of food and health products.