26 ISOLATION III 



is incomplete, the eluting solvent, usually methanol, can be removed by shaking 

 with water and the remaining solution dried, concentrated and again subjected 

 to chromatography. 



Adsorbents. The following materials, in finely divided form, have been used 

 for the chromatographic separation of carotenoids: alumina. Fuller's earth, 

 calcium carbonate, calcium hydroxide, kaolin, kieselguhr, magnesium oxide, 

 talcum, zinc carbonate, norite A, and others. The following 4 adsorbents are, 

 however, adequate for most purposes: aluminium oxide*, calcium hydroxide, 

 calcium carbonate, and zinc carbonate. Using these adsorbents it should be 

 possible to carry out successfully all chromatographic separations of carotenoids. 

 Alumina AlgOg: Suitable for the separation of carotenoid hydrocarbons, but 

 its use has recently decreased owing to its high cost. 



Calcium hydroxide Ca(0H)2: Calcium hydroxide was introduced for the 

 separation of carotenoid hydrocarbons by Karrer and Walker' in 1933 and 

 has since become the adsorbent most widely used for this purpose. It is cheap 

 and allows a complete separation of the epiphasic carotenoids. 



Calcium carbonate CaCOg : Calcium carbonate was iirst employed by Tswett 

 for the separation of carotenoids. Since 1 931, it has frequently been used for the 

 separation of phytoxanthins. 



Zinc carbonate ZnCOa: Zinc carbonate has frequently been employed, 

 especially within recent years. It was introduced by Karrer and is very 

 suitable for the separation of phytoxanthins. These are adsorbed somewhat 

 more strongly on zinc carbonate than on calcium carbonate. 



The following is the sequence of adsorbents of decreasing activity : Alumina, 

 aluminium hydroxide, magnesium oxide, calcium oxide, calcium hydroxide, zinc 

 carbonate, calcium carbonate, calcium sulphate, calcium phosphate, talcum, 

 sucrose, inulin'. 



Solvents. The purity of the solvents used is of the greatest importance for 

 the success of a chromatogram. The solvents must be dry and free from 

 impurities, such as alcohol, pyridine, sulphur-containing compounds, etc. For 

 the chromatography of epiphasic carotenoids, petroleum ether (b.p. 70-80°) or 

 a mixture of petroleum ether with benzene or ether are commonly used. 

 Recently petroleum ether-acetone mixtures have frequently been employed** 



For hypophasic carotenoids it is usual to employ benzene or a mixture of 

 benzene and ether or petroleum ether. Other solvents such as carbon di- 

 sulphide, ethyl acetate, etc. are also used, although the first-named solvents 

 are adequate in most cases. The following sequence of solvents, which is taken 



Different standardised grades of alumina are commercially available. For alumina 

 according to H. Brockmann, cf. Neuere Methoden der praparativen organischen Chemie, 



1943. P- 553- 



** See p. 42, reference 10. 



References p. 28. 



