Concentrations of serum phosphate and parathyroid hormone (PTH) are well known regulators of the production of 1,25-dihydroxyvitamin D (1,25-(OH)2D) and acidosis is known to affect the serum concentration of 1,25-(OH)2D. However, the factors that play a role in the regulation of serum 1,25-(OH)2D concentration in healthy subjects have not been fully evaluated. The associations of ionised calcium, pH, serum concentration of phosphate, PTH, 25-hydroxyvitamin D (25-OHD) and serum 1,25-(OH)2D were examined in 296 healthy premenopausal women (age range 17-40 years). Calculation of partial correlation coefficients showed that serum 1,25-(OH)2D was significantly correlated with phosphate (r = -0.148, P < 0.01), pH (r = 0.221, P < 0.001) and PTH (r = 0.136. P < 0.01). Ionised calcium was not related to serum 1,25-(OH)2D. When the results were stratified according to quartiles based on serum 1,25-(OH)2D concentration, significant trends (by analysis of variance) were seen in phosphate, pH, age, albumin and 25-OHD. Stepwise multiple regression analysis showed that phosphate and pH were the major contributors of serum 1,25-(OH)2D levels. There was a small contribution from PTH and 25-OHD. The results suggest that in young healthy premenopausal women plasma phosphate and pH may be important determinants of serum 1,25-(OH)2D concentration.