Measurement of circulating vitamin D in man

Abstract

An assay for vitamin D consisting of high-pressure liquid chromatography (HPLC) and ultraviolet (UV) absorbance detection has been developed and used to measure circulating vitamin D concentrations in human subjects during summer and winter and after deliberate exposure to ultraviolet radiation. Extracts of 2-4 ml of serum were initially fractionated on silica Sep-Pak cartridges followed by reverse-phase HPLC and finally quantitated by UV-absorbance during straight-phase HPLC. Using these methods, we determined the normal range for circulating vitamin D in Boston subjects to be less than 0.5 ng/ml to 25 ng/ml (n = 30); subjects sampled during summer months had higher concentrations of vitamin D than those sampled during winter months. In subjects exposed to a single quantitative dose of ultraviolet radiation (UVR), large transient increases in circulating vitamin D3 were observed. Concentrations rose 30-50 fold over the first days after exposure before returning to basal levels by one week.