Phosphate is important for several metabolic functions and essential for bone mineralization. Sex-differences exist in the relation between serum phosphate and certain diseases. The reference interval for phosphate is age-adjusted in infants, but most institutions use the same intervals for adult men and women despite increasing evidence for age and sex-differences. We aimed to study these differences in two large population-based cohorts in order to evaluate whether current reference intervals are adequate. 8837 participants from three cohorts of the Rotterdam Study (RS) and 422,443 participants from UK Biobank (UKBB), aged 40 and older and without chronic kidney disease, were analyzed for sex/differences in serum phosphate using standard reference values (0.8-1.45 mmol or 2.5-4.5 mg/dL). Analyses were further stratified in women by menopausal status. Women had higher serum phosphate concentrations and a higher population range compared to men in all cohorts. Hypophosphatemia was more prevalent in men and hyperphosphatemia was more prevalent in women. Sex-differences were present in all age-categories. Perimenopausal women had higher serum phosphate concentrations than men of the same age, but lower than postmenopausal women of the same age. This study in two population-based cohorts showed that women have higher serum phosphate concentrations than men and that women show a marked increase in serum phosphate during menopause. Moreover, the population range for serum phosphate was higher in women than in men. These findings indicate a need for sex-specific reference intervals for serum phosphate in adults older than 45 years.