Diabetes mellitus is associated with a decrease in bone mineral content and increased urinary excretion of calcium and phosphate. The purpose here was to elucidate the pathogenesis of these changes by comparing serum calcium fractions of diabetics and healthy controls. In a cross-sectional study, serum ionized calcium at pH 7.40, [Ca2+]7.4, of 46 insulin-treated diabetics was decreased in comparison with 44 healthy controls: mean 1.16 (SD 0.04) vs 1.21 (SD 0.03) mmol/L (p less than 0.001). The decreased [Ca2+]7.4 was associated with a higher concentration of complexed calcium in diabetic serum. The diabetics had higher concentrations of undetermined anions (p less tha 0.001) and this anion gap correlated negatively with [Ca2+]7.4 (r = -0.43, p less than 0.02). Serum [Mg2+] was decreased in the diabetics (p less than 0.001). Values for venous acid-base status, serum creatinine, total and ultrafiltrable calcium, parathyrin, and inorganic phosphate were the same in the two groups. [Ca2+]7.4, but not serum total calcium, had increased by 0.04 mmol/L (p less than 0.001) in the diabetics and by 0.02 mmol/L (p less than 0.01) in the controls by 90 min after breakfast, with and without subcutaneous insulin, respectively.