24,26-Dihydroxyvitamin D2: a unique physiological metabolite of vitamin D2

A new vitamin D2 metabolite, 24,26-dihydroxyvitamin D2, has been detected in the plasma of rats fed physiologic amounts of vitamin D2. The identity of the new metabolite (isolated from cow plasma) was established by ultraviolet absorbance, mass spectroscopy, chemical reactivity, and NMR spectroscopy. Among these, the mass spectrum was unique for the presence of a peak at M-48 that was attributed to an intramolecular rearrangement involving both the C-24 and C-26 hydroxyl groups. A 300-MHz 1H NMR spectrum of 40 micrograms of metabolite indicated a downfield shift of the C-28 methyl group signal to delta 1.30 and a multiplet at delta 3.66 corresponding to the hydroxylated C-26 methyl group. We determined that the formation of 24,26-dihydroxyvitamin D2 represented a major pathway for further metabolism of 24-hydroxyvitamin D2 in rats, exceeding the formation of 24,25-dihydroxyvitamin D2. Standard bioassays revealed that 24,26-dihydroxyvitamin D2 possessed very little biological activity and most likely represents a deactivation pathway for 24-hydroxyvitamin D2.