Measurement of 25-hydroxyvitamin D in serum and its relation to sunshine, age and vitamin D intake in the Danish population

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Abstract
A competitive protein-binding assay for 25-hydroxyvitamin D (25-OHD) based upon a specific binding protein in the cytosol from rachitic rat kidneys is described. A diethyl ether extraction followed by separation by freezing was used. The extracts were chromatographed on short silicic acid columns, which separated 25-hydroxycholecalciferol from cholecalciferol, 24,25-dihydroxycholecalciferol, and 1,25-dihydroxycholecalciferol. A small aliquot of the 25-OHD fraction was used in the assay and free and bound vitamin were separated by dextran coated charcoal. The lower detection limit was 0.8 ng/ml (2.0 nmol/1). The levels of 25-OHD were measured in 596 healthy subjects and a seasonal variation was demonstrated. This variation, however, was only found in those without regular vitamin D intake, whereas the level of 25-OHD remained constant throughout the year in subjects with regular vitamin D supplement. The levels of 25-OHD were lower in the elderly subjects compared to younger ones, but seasonal variation was observed in both groups. In the summer months there was a significant correlation between age and the 25-OHD level. The mean levels of 25-OHD in Denmark are within the range of means found in the United States and Sweden but are higher than those reported from England, Belgium and France. This indicates a relatively high vitamin D intake in the Danish population and a low risk of nutritional vitamin D deficiency.