238 PIGMENTS 



EXTRACTION OF CHLOROPHYLL. 



The usual method of extracting chlorophyll from green 

 tissues consists in first steeping the fresh material in hot water 

 to destroy oxidizing enzymes and then extracting the colour- 

 ing matter by means of warm alcohol. Willstatter, however, 

 recommends the use of dried in place of fresh material, and 

 extracting by shaking with organic solvents (ethyl or methyl 

 alcohol, ether or acetone) in the cold. 



The chief advantage in using dried material lies in the 

 fact of its relatively small bulk, lOO grams of stinging nettle 

 leaves, for example, weighing only 25 grams after drying. It 

 has been shown, moreover, that the operation of drying pro- 

 duces no change of any importance in the chlorophyll, since 

 the results obtained from dried material have been repeated 

 and confirmed on fresh material. 



On the other hand, organic solvents containing an appreci- 

 able amount of water are preferable to the dry solvents. This 

 is attributed by Willstatter to the fact that aqueous solvents 

 dissolve out salts, such as potassium nitrate, from the cell sap, 

 and these affect the state* of the colloidal solution of chloro- 

 phyll in the chloroplast, thereby rendering the chlorophyll 

 more easily accessible to the solvent. Moreover, the number 

 of substances going into solution is thereby increased, and the 

 solution is no longer effected by the solvent alone but by the 

 solvent together with the accessory substances. 



If dry solvents are used, the extract is much less pure since 

 it contains a larger proportion of carotinoids, lecithins, etc, 

 whose solubilities are very similar to those of chlorophyll. 



The following methods of extracting dried or fresh leaves 

 respectively are described by Willstatter : — 



I. Half a kilo of dried material is spread on a porcelain 

 Buchner funnel in a layer of not more than 4 to 5 cms. thick, 

 and I '5 litres of solvent are drawn through this layer by means 

 of a filter pump in the course of half an hour. This filtrate, 

 measuring about 0*9 litre, contains from 4-25 to 4-5 grams of 

 chlorophyll. 



The solvent employed may be either 90 per cent (aqueous) 

 alcohol or 80 per cent (aqueous) acetone. The former filters 



*See section on Colloids, page 283. 



