This study was conducted to examine the effect of calcium intake on the rise in serum 25-hydroxyvitamin D [25(OH)D] levels in response to supplemental vitamin D(3). Fifty-two healthy older men and women were randomly assigned to take calcium (500 mg twice daily with meals) or placebo tablets for 90 d between October 1 and the end of March. All participants were placed on 800 IU/d (20 microg/d) vitamin D(3). Serum 25(OH)D measurements were made at baseline and on d 30, 60, and 90. The mean baseline 25(OH)D values were 19.2 +/- 6.4 ng/ml (47.9 +/- 15.9 nmol/liter) in the calcium group and 19.6 +/- 6.7 ng/ml (49.1 +/- 16.7 nmol/liter) in the control group (P = 0.808). The difference in pattern of change in 25(OH)D was not statistically significant (group by time interaction, P = 0.651); the calcium group increased 6.5 +/- 5.9 ng/ml (16.2 +/- 14.8 nmol/liter; P < 0.001), and the control group increased 6.6 +/- 7.0 ng/ml (16.6 +/- 17.4 nmol/liter; P < 0.001). The 95% confidence interval for difference in mean increase, calcium vs. control, was -3.8 +/- 3.5 ng/ml (-9.6, 8.7) nmol/liter. In older men and women, the level of calcium intake, within the range of 500-1500 mg/d, does not have an important effect on the rise in serum 25(OH)D that occurs in response to 800 IU (20 microg)/d vitamin D(3).