Of a series of our works, this report is concerened to the chromatographical analysis of vitamin concentrate. The purpose of the chromatography is to clarify the nature of the concentrate and obtain the data necessary for the further concentration. The followings are the outlines of our experiment.
I) Vitamin concentrates obtained by the saponification method were divided into three fractions by the use of chromatographical technique. The qualities and the contents of nonvitamin substances in the fractions were examined (Tables 1, 2, 5 and Figs. 2, 4, 5).
The chromatographic column used, is shown in Fig. 1. As adsorbents, Merk's alumina weakened by 3.3% methanol is used. Dissolve 0.3 gr. of unsaponifiable matter in light petroleum to pass through the column and then elute by different solvents into the following three fractions.
Fraction I is developed with light petroleum to make the total volume of solvent 50m
l. It contains anhydrovitamin A and other hydrocarbons.
Fraction II is developed with benzene containing 5% methanol, to make the total volume of solvent 50m
l, and contains vitamins, sterols, and other mono-, and dihydric-alcohols.
Fraction III is the residue remaining in the column.
II) Maleic anhydride reacts rapidly on conjugated double bonds in vitamin A and other compounds in unsaponifiable matter. Samples of concentrates were treated with maleic anhydride in benzene for six hours on boiling water bath. After saponified, unsaponifiable matter was extrated with ether. The extracts having no conjugated double bonds, were also divided into three fractions by the chromatographical method mentioned above (Tables 3 and 6).
After comparing the result with that of original concentrates (Tables 1 and 5), the weight percent diagrams of vitamin concentrates were obtained (Fig. 6).
III) The same chromatographical analysis was applied to each molecular distillate of vitamin concentrate (Tables 4, 5, 6, and Fig. 3). From the result, it was ascertained that molecular distillation was very useful to refine the concentrate and to remove non vitamin materials, having conjugated double bonds. But the method is not applicable to removing compounds with no conjugated double bonds, for the distillate contains the compounds as much as original concentrate (Figs. 6 and 7).
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