344 PHOTOSYNTHESIS 



The yellozv pigments carotin mid xanthophyll are widely distributed 

 in nature, in both plants and animals, and as they apparently occur 

 in different isomorphous and isomeric modifications these have, at the 

 suggestion of Tswett, been grouped under the term carotinoids. We shall 

 here confine the discussion to the yellow pigments in the chloroplasts 

 of green leaves and not include the pigments of "autumn leaves" nor of 

 yellow, orange and red flowers and fruits. 



Arnaud ® succeeded in isolating carotin from green leaves by extrac- 

 tion with petroleum ether. Leaves of spinach, vacuum dried and powdered, 

 were extracted with petroleum ether. The petroleum ether was distilled 

 and from the concentrated solution the solvent was evaporated at ordi- 

 nary temperature. The waxy substances were removed by washing the 

 residue with a little cold ether and the pigment was recrystallized from 

 petroleum ether. When the leaf material is perfectly dry rapid extraction 

 with petroleum ether removes no chlorophyll nor xanthophyll. 



Willstatter obtained the yellow pigments as a side product in the 

 preparation of chlorophyll. The plant material was extracted with 80 

 per cent acetone and the dissolved pigments were transferred to petroleum 

 ether, as has already been described. The petroleum ether solution is freed 

 from xanthophyll by washing three times with 80 per cent methyl alco- 

 hol. These extracts are bright green. The pigment is extracted from 

 the methyl alcohol by the addition of ether (4 to 5 liters) and dilution 

 with water. The ether solution contains xanthophyll and some chloro- 

 phyll b. This latter can be removed by shaking the ether solution with 

 30-50 cc. of concentrated methyl alcoholic potassium hydroxide and sub- 

 sequent extraction with water. Direct crystallization of the xanthophyll, 

 without removal of the chlorophyll b, can be accomplished by drying 

 the ether with sodium sulfate, concentrating to about 30 cc. and adding 

 200-300 cc. methyl alcohol. This solution is again heated gently to 

 evaporate the ether and is filtered. On cooling the xanthophyll crystal- 

 lizes in shining plates. In order to have the xanthophyll crystallize out 

 more completely water may be added to the methyl alcohol solution ; under 

 these conditions the xanthophyll separates in crystal aggregates which 

 gradually change into plates. The yield of xanthophyll is about 0.8 gram 

 for 2 kg. of dried nettle leaves. 



The petroleum ether chlorophyll solution from which the xanthophyll 

 has been removed with 80 per cent methyl alcohol, as just described, 

 still contains carotin. The petroleum ether solution is washed four times, 

 each with 2000 cc. of water. Thus the methyl alcohol is removed and 

 the chlorophyll is precipitated. The petroleum ether, after filtration, is 

 concentrated in vacuum at 40° and the oily residue is treated with 300 

 cc. 95 per cent alcohol. The carotin rapidly separates out in steel-blue 

 crystals. In order to free the carotin from impurities it is dissolved in 



•Arnaud, Compt. rend., 100, 751 (1885). Van Wisselingh, Flora, 107, 371 

 (1915). Coward, Biochem. Jour., 18, 1114, 1123 (1924). 



