26-Hydroxylation of 1α,25-dihydroxyvitamin D3 does not occur under physiological conditions

The 26-hydroxylation of 1 alpha ,25-dihydroxyvitamin D-3 in rats in vitro and in vivo was studied under physiological conditions. Incubation of 1 alpha ,25-dihydroxy-[26,27-H-3]vitamin D-3 with rat kidney or rat liver homogenate showed formation of a metabolite that was identified as 1 alpha ,25(5),26-trihydroxy-[26,27-H-3]vitamin D-3 by comigration on three different HPLC systems and a periodate cleavage reaction. This metabolite was not generated by hydroxylation of 1 alpha ,25-dihydroxy-[26,27-H-3]vitamin D-3 itself but by an enzymatic conversion of a precursor that was formed nonenzymatically in substantial amounts upon storage of 1 alpha ,25-dihydroxy-[26,27-H-3]vitamin D-3 in ethanol at -20 degreesC under argon for more than three weeks. An in vivo metabolism study in rats dosed with a physiological dose of 1 alpha ,25-dihydroxy-[26,27-H-3]vitamin D-3 confirmed the absence of 26-hydroxylation of the hormone. As expected at 6 h postinjection of purified 1 alpha ,25-dihydroxy-[26,27-H-3]vitamin D-3, 1 alpha ,24(R),25-trihydroxy-[26,27-H-3]vitamin D-3, as well as traces of (23S,25R)-1 alpha ,25-dihydroxy-[H-3]vitamin D-3-lactone were detected and identified on straight phase and reverse phase HPLC in serum, kidney, and liver, (C) 2001 Academic Press.