<p>Fatigue is induced by physical and mental overload and decreases activity, motivation, and judgment, interfering with daily life. An association between fatigue and prolonged activation of the immune system has been suggested. However, the detailed underlying mechanisms are unknown, and effective treatments for fatigue have not been established. Ninjin’yoeito, a Kampo medicine, reportedly improves anorexia and fatigue scores. In this study, we examined the anti-fatigue effects of Ninjin’yoeito extract powder (NYT) in a polyinosinic-polycytidylic acid (poly[I:C])-induced mouse model of inflammatory fatigue. C57BL/6 male mice were intraperitoneally injected with poly(I:C) (12 mg/kg) to assess locomotive activity and nesting and grooming behaviors. NYT was orally administered to mice. NYT showed no effect on the reduction of locomotive and wheel-running activities following poly(I:C) administration. The enhanced rectal temperature and peripheral and hypothalamic levels of inflammatory cytokines also remained unaffected by NYT. In contrast, NYT significantly restored the nesting and grooming behaviors impaired by poly(I:C) administration. The restorative effect of NYT was abolished by cotreatment with a dopamine D2 receptor (D2R) antagonist. Furthermore, the impaired grooming behavior following poly(I:C) administration was restored by the dopamine-metabolizing enzyme monoamine oxidase (MAO)-B inhibitor, but not by the MAO-A inhibitor. Ingredients of NYT, such as formononetin derived from the astragalus root, paeonol from the peony root, heptamethoxyflavone and nobiletin from citrus unshiu peel, demonstrated inhibitory activities against MAO-B. These findings suggest that NYT improves fatigue-like self-care deficits potentially by inhibiting dopamine metabolism, likely via MAO-B, leading to enhanced D2R signaling.</p> Graphical abstract <p></p>