Simple Laboratory Experiments on Leaf Pigments. 35 



Experiment 12. Microscopic examination of ethyl chlorophyllide. 



Prepare sections of fresh Heracleum leaves and mount them 

 in a drop of 90% alcohol. Leave the slide under a hell jar contain- 

 ing a dish of alcohol. The section slowly dries in the course of 

 half a day or a day. It is then examined under the microscope 

 when there will be observed the characteristic triangular and 

 hexagonal crystals of ethyl chlorophyllide (crystalline chlorophyll). 



Experiment 13. Production of methyl chlorophyllide in the leaf. 



Sections may be used as in the preceding experiment, or a 

 piece of a leaf may be employed. In the latter case a test-tube 

 with 4 c.c. 75% methyl alcohol is taken and 1 gram of fresh leaf is 

 added to it. The leaf first becomes a darker green and then during 

 the course of a few hours becomes yellowish. On holding the leaf 

 to the light there can be observed with the naked eye a number of 

 black points. If sections of the leaf be cut and examined under the 

 microscope, these spots appear as aggregates composed of rhom- 

 bohedral crystals, occurring only in certain cells. 



Experiment. 14. Extraction of ethyl chlorophyllide. 



Two grams of dry Heracleum leaf powder is left for a day in a 

 test-tube containing 6 c.c. 90% alcohol. The extract is then 

 filtered through a small Buchner funnel and the powder on the 

 filter washed with a little acetone. The filtrate is washed with the 

 same quantity of ether, and then with wal;er. The ether solution is 

 transferred to a separating funnel and washed with watei-, and then 

 concentrated on a water bath to ^ or 1 c.c, and 3 c.c. petrol ether 

 is added. On standing, the ethyl chlorophyllide is precipitated in 

 the form of crystalline aggregates. It is freed from yellow pigments 

 by shaking with a little ether, and can be further purified by 

 redissolving in ether and precipitating again with petrol ether. 



Experiment 15. Spectroscopic examination. Required: small 

 spectroscope and glass vessel with parallel sides of about 1 cm. in 

 width ; source of light (incandescent burner or Nernst lamp or 

 sunlight). 



The following absorption spectra may be examined : — 

 a. Chlorophyll spectrum from acetone extract obtained in Experiment 1 . 

 The extract is diluted with about five times its volume of 85% 

 acetone. The spectrum shows a main absorption in the red at the 

 Praunhofer line C. Then follow, towards the violet, three absorp- 

 tion bands decreasing in intensity, and the end absorption in the 



