Background: Uncertainty remains whether ageing before late adulthood and menopause reduce fat-free mass and fat mass-adjusted resting energy expenditure (REEadj). We therefore investigated whether REEadj differs 1) between middle-aged and younger women and 2) in middle-age by menopause status. We repeated the age group comparison between middle-aged mothers and their daughters to partially control for genotype. We also explored whether oestradiol and follicle-stimulating hormone concentrations or body fat percentage explain REEadj in midlife. Methods: We divided 120 women, including 16 mother–daughter pairs, into age groups. Group I (n=26) consisted of participants aged 17-21, group II (n=35) aged 22–38 and group III (n=59) aged 41-58 years. Group III women were pre- or perimenopausal (n=19), postmenopausal (n=30) or postmenopausal hormone therapy users (n=10). REE was assessed using indirect calorimetry, body composition using dual-energy X-ray absorptiometry and hormones using immunoassays. Results: Group I had 126 kcal/d (95% confidence interval [CI], 93–160; P<0.001) higher REEadj than group III, while group II had 88 kcal/d (CI, 49-127; P<0.001) higher REEadj. Furthermore, the daughters had 100 kcal/d (CI, 63-138; P<0.001) higher REEadj than their mothers. In group III, REEadj did not differ by menopause status, but body fat percentage was positively associated with REEadj (beta=0.39, P=0.003). Conclusions: We demonstrated that REE declines with age in women before late adulthood, even when controlling for body composition and partially for genetic background, and that menopause does not contribute to the decline in REEadj. Menopause-associated fat accumulation may actually elevate REEadj.