Barley intake reportedly reduces the risk of cardiovascular disease, but effects on the systemic phenotypes during healthy aging have not yet been examined. Therefore, we examined the effects of barley on the lifespan; behavioral phenotypes, such as locomotor activity, and cognitive functions, and intestinal microbiome in the senescence-accelerated mouse-prone 8 (SAMP8) mouse. We prepared two mild high-fat diets by adding lard, in which the starch components of AIN-93G were replaced by rice or barley "Motchiriboshi." SAMP8 (four weeks old, male) mice were fed AIN-93G until eight weeks old, and then rice (rice group) or barley diet (rice: barley = 1:4, barley group) until death. Changes in aging-related phenotypes, object and spatial recognition, locomotor and balancing activities, and the intestinal microbiome were recorded. Moreover, plasma cholesterol levels were analyzed at 16 weeks old. Barley intake prolonged the lifespan by approximately four weeks, delayed locomotor atrophy, and reduced balancing ability and spatial recognition. Barley intake significantly increased the medium and small particle sizes of high-density lipoprotein (HDL) cholesterol, which is associated with a reduced risk of total stroke. The Bacteroidetes to Firmicutes ratio in the barley group was significantly higher than that in the rice group during aging. Thus, lifelong barley intake may have positive effects on healthy aging.