A new chromatographic system for vitamin D3 and its metabolites: resoluation of a new vitamin D3 metabolite.

A simple yet powerful new chromatographic procedure for vitamin D(3) and its metabolites is described. Liquid-gel partition chromatography on Sephadex LH-20 using a solvent of various percentages of CHCl(3) in Skellysolve B (petroleum ether, bp 67-69 degrees C) permits excellent resolution of vitamin D(3), 25-hydroxyvitamin D(3), and their more polar metabolites. Of special importance is the resolution of the metabolites of vitamin D(3) more polar than 25-hydroxycholecalciferol. Because of this resolution, a new metabolite of vitamin D(3) has been demonstrated in the plasma of rats and in the intestines of chicks given 100 IU of vitamin D(3)-1,2-(3)H.