title: 25-Hydroxylase activity in subcellular fractions from human liver. Evidence for different rates of mitochondrial hydroxylation of vitamin D2 and D3.
abstract: 25-Hydroxylation of vitamin D2 and D3 was studied in subcellular fractions from human liver, using a technique based on isotope dilution-mass spectrometry. The mitochondrial fraction fortified with isocitrate catalysed 25-hydroxylation of vitamin D3 at a rate of about 10 pmol/mg protein X min. Under the same conditions, the rate of 25-hydroxylation of vitamin D2 was less than 2 pmol/mg protein X min. Crude microsomes fortified with NADPH catalysed 25-hydroxylation of vitamin D3 to a very low extent, and this activity was not linear with the amount of microsomal protein. A higher rate of conversion was obtained with a partially purified cytochrome P-450 fraction in the presence of NADPH-cytochrome P-450 reductase and NADPH. This fraction also catalysed 25-hydroxylation of 1 alpha-hydroxyvitamin D3 and 5 beta-cholestane-3 alpha, 7 alpha, 12 alpha-triol. 25-Hydroxylation of vitamin D2 could not be detected, neither with crude microsomes, nor with the microsomal cytochrome P-450 fraction. Since the assay for 25-hydroxyvitamin D2 was less sensitive than that for 25-hydroxyvitamin D3, these experiments do not rule out the presence of some 25-hydroxylase activity towards vitamin D2 in the microsomes. The results are discussed in relation to previous work in which a lower toxicity has been reported for vitamin D2 than for vitamin D3 in some mammalian species.
mesh_terms: D002762:Cholecalciferol / Q000378:metabolism*; D003577:Cytochrome P-450 Enzyme System / Q000378:metabolism; D004872:Ergocalciferols / Q000378:metabolism*; D006801:Humans; D006900:Hydroxylation; D013058:Mass Spectrometry; D008862:Microsomes, Liver / Q000378:metabolism*; D008930:Mitochondria, Liver / Q000201:enzymology*; D006899:Mixed Function Oxygenases / Q000378:metabolism*
chemical_list: D004872:Ergocalciferols; D002762:Cholecalciferol; D003577:Cytochrome P-450 Enzyme System; D006899:Mixed Function Oxygenases