Studies on the biogenesis of vitamin D. I. Identification and determination of 7-dehydrocholesterol in rat skin.

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Abstract
Lipids were extracted from the skin of normal rats with chloroform-methanol (1:1) and the solvent evaporated. The resulting residue was saponified, and the unsaponifiable matter extracted with benzene was purified by subjecting the extract successively to hydroxyalkoxypropyl (HAP) Sephadex column chromatography, silica-gel column chromatography and preparative TLC. The resulting purified provitamin D fraction contained 7-dehydrocholesterol (7-DHC) based on data of UV spectrum, high-performance liquid chromatography (HPLC), GLC and GLC-mass spectrometry (MS). Based on the results of GLC-MS and mass chromatography, the fraction contained small amounts of cholesterol, lathosterol and 2 other unknown sterols besides 7-DHC. The determination of 7-DHC in rat skin was then investigated, and an HPLC method was proposed. This method included extraction of lipids from small pieces of rat skin with chloroform-methanol (1:1), saponification, isolation of the unsaponifiable matter and application to HPLC. The estimated average values of 7-DHC in normal rat skin (20 wk old), with and without UV-irradiation were 107.7 and 103.8 .mu.g/g, respectively. These were significantly higher than the corresponding values of vitamin D-deficient rat skin. UV-irradiation of rat skin did not affect the change of 7-DHC content in the tissue.

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