The retention behavior of N-acetylglucosaminides of vitamin-D2and -D3, and those of provitamin-D2and -D3are examined using reverse phase high performance liquid chromatography. Inclusion chromatography using cyclodextrin as the mobile phase additive is also used for this purpose. The addition of methyl-β-cyclodextrin to the mobile phase is effective in separating the pair of N-acetylglucosaminides of vitamin-D2and -D3or those of provitamin-D2and -D3.