Numerous studies from our and other laboratories have shown that women have a lower respiratory exchange ratio (RER) during exercise than equally trained men, indicating a greater reliance on fat oxidation. Differences in estrogen concentration between men and women likely play a role in this sex difference. Differing estrogen and progesterone concentrations during the follicular (FP) and luteal (LP) phases of the female menstrual cycle suggest that fuel use may also vary between phases. The purpose of the current study was to determine the effect of menstrual cycle phase and sex upon glucose turnover and muscle glycogen utilization during endurance exercise. Healthy, recreationally active young women ( n = 13) and men ( n = 11) underwent a primed constant infusion of [6,6-2H]glucose with muscle biopsies taken before and after a 90-min cycling bout at 65% peak O2 consumption. LP women had lower glucose rate of appearance (Ra, P = 0.03), rate of disappearance (Rd, P = 0.03), and metabolic clearance rate (MCR, P = 0.04) at 90 min of exercise and lower proglycogen ( P = 0.04), macroglycogen ( P = 0.04), and total glycogen ( P = 0.02) utilization during exercise compared with FP women. Men had a higher RER ( P = 0.02), glucose Ra ( P = 0.03), Rd ( P = 0.03), and MCR ( P = 0.01) during exercise compared with FP women, and men had a higher RER at 75 and 90 min of exercise ( P = 0.04), glucose Ra ( P = 0.01), Rd ( P = 0.01), and MCR ( P = 0.001) and a greater PG utilization ( P = 0.05) compared with LP women. We conclude that sex, and to a lesser extent menstrual cycle, influence glucose turnover and glycogen utilization during moderate-intensity endurance exercise.