Simple Laboratory Experiments on Leaf Pigments. 27 



Two grams of leaf powder are sucked to a filter paper on the 

 Buchner funnel and a small quantity of the solvent added. This is 

 allowed to soak into the powder for a few minutes. The fluid is 

 then sucked through with the pump. The operation is repeated until 

 all the 20 c.c. of solvent has been added, when the powder is sucked 

 dry. A deep blue green solution, with red fluorescence, is obtained 

 which contains all the four pigments from the leaf. Usually the 

 powder will still be coloured green as the extraction is not generally 

 complete. 



Experiment 2. Transfer of the pigments from an acetone solution 

 to an e ther, iff t» a petrol ether, solution. Required : 1 separating 

 funnel ; about 10 c.c. ether and 10 c.c. petrol ether; 5 c.c. acetone 

 extract of leaves. 



Five c.c. of the acetone extract obtained in Experiment 1 are 

 poured into double the quantity of ether contained in a separating 

 funnel. An equal quantity of distilled water is added, this being 

 poured gently down the side of the funnel in order to avoid the 

 formation of emulsions. In the course of a few minutes, the ether 

 layer separates out and now contains the pigments. The lower 

 layer, which is slightly green, is run off. The addition of distilled 

 water and subsequent removal of the lower layer is repeated about 

 four times, in order completely to remove the acetone from the 

 ether solution. If the ether solution should have become at all 

 emulsifled, it can be cleared by shaking with anhydrous sodium 

 sulphate and Altering. 



A petrol ether solution may be obtained in the same way by 

 using 10 c.c. of petrol ether in place of ether. 



Experiment 3. Demonstration of the two green pigments. 

 Required : 10 c.c. petrol ether solution of mixed pigments ; 10 c.c. 

 92% methyl alcohol ; 2 separating funnels. 



The petrol ether solution from the last experiment is shaken 

 with 10 c.c. 92% methyl alcohol. Two layers are formed of which 

 the petrol ether layer contains chlorophyll a and the methyl alcohol 

 layer chlorophyll b. The solution of chlorophyll a is blue green 

 while that of chlorophyll b is a purer green, but the colour differ- 

 ence between them is diminished owing to the presence of the 

 yellow pigments, of which carotin is in the petrol ether, and 

 xanthophyll in the methyl alcohol. 



A characteristic difference between the two green pigments is to 

 be found in the phase which appears on saponification with methyl 



