32 CAROTINOIDS AND RELATED PIGMENTS 



yellow. Kraus named the green pigment cyanophyll and the yellow 

 pigment xanthophyll. From our present knowledge of the relative 

 solubility properties of the carotinoids which have been built very 

 largely upon Kraus' observations, it is obvious that his xanthophyll 

 pigment was composed almost entirely of the xanthophyll carotinoids, 

 although traces of carotin may have been present also. Kraus found 

 that the absorption bands of his xanthophyll fraction in the blue part 

 of the spectrum corresponded with Bands V and VI of the leaf extract. 

 The residue from Kraus' xanthophyll fraction gave a deep blue colora- 

 tion with concentrated sulphuric acid, and the xanthophyll solution 

 itself bleached very quickly in the sunlight. 



Several studies of the leaf pigment, using the Kraus procedure, soon 

 followeH. Konrad (1872) observed that the separation of xanthophyll 

 from chlorophyll by benzene was effective only when an alcohol of 

 about 70 per cent (by volume) strength was employed. Treub (1874) 

 substantiated the necessity of using weaker alcohol for the benzene 

 separation, and found that CS, was effective when the chlorophyll 

 extract was in strong alcohol. Ccmpert (1872) found that linseed oil 

 could be used in place of benzene and Wiesner (1874a, b) found that 

 a number of vegetable oils (linseed, walnut, poppyseed, olive) and 

 ethereal oils (turpentine oil, rosemary oil, oil of wintergreen) and also 

 carbon disulfidc could be employed. Wiesner used 85 per cent (by 

 volume) alcohol and benzene, boiling at 92-94 C., for the regular 

 Kraus separation. Equally good results were obtained with toluene 

 and xylene, or mixtures of these with benzene. Wiesner even suc- 

 ceeded in shaking the green chlorophyll out of alcoholic leaf extracts 

 with dilute egg albumin. Especially important was his observation 

 that dilute ammonia or caustic alkali solutions acting on the residues 

 from alcoholic leaf extracts would take up most of the yellow color 

 leaving behind the chlorophyll, mixed with a little xanthophyll. This 

 observation, pointing to the resistance of the yellow chromolipoids to 

 alkalis and the attacking;; of chlorophyll by the same reagents, was 

 later developed by Hansen (1884a) and is still widely used for the 

 separation of carotinoids from chlorophyll. Hansen boiled young 

 wheat plants in water for one-half hour, dried the product, extracted 

 with cold 96 per cent alcohol in the dark, concentrated the extract to 

 one-eighth volume, saponified with NaOH, diluted the soap solution 

 with water, and salted out the soap with NaCl. The green soap thus 

 obtained was extracted with petroleum ether, which gave a yellow 

 extract which Hansen called "chlorophyll gelb," the residue being 



