Milk fat globule membranes (MFGMs) are complex structures that incorporate bioactive proteins and lipids to assist in infant development. However, the antifatigue and antioxidant potentials of MFGM have not been investigated. In this study, repeated force swimming measured fatigue in male BALB/c mice fed MFGM and saline for 18 weeks. The MFGM supplementation increased the time to exhaustion by 42.7% at 6 weeks and 30.6% at 14 weeks (p p p p Bacteroides, Butyricimonas, and Anaerostipes. Our results demonstrate that MFGM may maintain redox homeostasis to relieve fatigue, suggesting the potential application of MFGM as an antifatigue and antioxidant dietary supplement.