To elucidate the age-related sex difference in glucose tolerance, we conducted 75 g oral glucose tolerance tests in 1156 participants. Participants were divided into four groups, namely, young (22-29) males, young females, middle-aged (>50) males, and middle-aged females. According to the Japanese Clinical Practice Guideline for Diabetes 2019, the prevalence of normal glucose tolerance (NGT) was significantly lower in middle-aged than in young participants. The prevalence of high-normal fasting plasma glucose (FPG) was higher, and NGT was lower in young males (high-normal FPG 15.2%, NGT 82.0%) than young females (high-FPG 3.9%, NGT 94.3%). Combined glucose intolerance (CGI) was higher and NGT was lower in middle-aged males (CGI 10.2%, NGT 25.2%) than in middle-aged females (CGI 3.3%, NGT 39.8%). FPG and body mass index (BMI) were the lowest and Homeostatic model assessment beta cell function (HOMA-β) was the highest in young females, followed by young males, middle-aged females, and middle-aged males. Multiple linear regression analysis revealed that BMI weakly correlated with HOMA-β and Matsuda index in all subjects except young females. The superior glucose tolerance in females was apparent in young, but attenuated in middle-aged females. The differences are due to the higher insulin secretion potential and lower BMI in young females.