Isolation and identification of new constituents in milk fat

After removal of cholesterol on a digitonin column, the unsaponifiable matter of milk fat was examined for alcoholic substituents. Derivatization with pyruvic acid chloride 2,6-dinitrophenylhydrazone and fractionation of the derivatives gave four main fractions. The second, the hexane-benzene fraction, was shown by thin-layer chromatography to have a mobility similar to many common sterols. The hexane-benzene fraction was saponified and gave rise to free alcohols, which were then analyzed on a combination gas-liquid chromatograph-mass spectrometer. Dihydrolanosterol, previously unreported in milk fat, and lanosterol, previously identified but never confirmed, were characterized. The sterols which were precipitated on the digitonin column, were removed, and by the use of the combination gas-liquid chromatograph-mass spectrometer beta-sitosterol was identified. In addition, lanosterol and dihydrolanosterol were isolated from the unsaponifiable matter by chromatography on Florisil.