526 PRACTICAL ORGANIC AND BIO-CHEMISTRY 



The chlorophyll mixture (8 gtn.) is dissolved in 150-200 c.c. of ether, 

 filtered and poured into 4000 c.c. of petrol ether. By adding 50-100 c.c. 

 of methyl alcohol any precipitate is dissolved. The solution is freed from 

 ether by shaking twice with 2 litres of 80 per cent, methyl alcohol. This 

 serves also to remove the yellow pigments if raw chlorophyll be used. 



The petrol ether is shaken 14-16 times with 2 litres of 85 per cent, 

 methyl alcohol which has been saturated with petrol ether (5*5-10 per 

 cent, are taken up) and containing -01 gm. of oxalic acid per litre. 



The petroleum ether layer containing the chlorophyll a is washed with 

 water which throws down the pigment. It is collected upon 30-100 gm. 

 of talc, filtered off on talc and washed with petrol ether. The pigment is 

 dissolved by shaking the talc in a bottle with ether, the blue solution is 

 filtered repeatedly and concentrated ; finally the ether is removed in a 

 desiccator until a dry mass is obtained. 



The first methyl alcohol extract containing the chlorophyll b is diluted with 

 i litre of methyl alcohol, washed with i litre of petrol ether, poured into 2 

 litres of ether and partition of the solvents effected by adding water. 



The second methyl alcohol extract is diluted and treated with the wash 

 petrol ether from the first extract + litre petrol ether more. It is transferred 

 to the ether from the first extract + i litre of ether more. 



In each case the petrol ether is freed from methyl alcohol with water. 

 The third and fourth extracts are treated in the same way. The other ex- 

 tracts are mixed with gradually decreasing quantities of methyl alcohol, 900, 

 800, 700 c.c., etc. They are washed with the same petrol ether increased 

 in volume by adding \ litre more. 



The pigment is in each case transferred from the methyl alcohol to ether by 

 adding water. The solvent consists of ether + petrol ether. It is freed from 

 methyl alcohol by washing with water, dried with sodium sulphate and evapor- 

 ated to 500 c.c. Further evaporation to 30 or 40 c.c. is carried out in vacua ; 

 the pigment is precipitated by adding 300 c.c. of petrol ether and filtered on talc ; 

 the filtrate contains chlorophyll a. It is purified by dissolving in ether and 

 precipitating with petrol ether several times ; it is washed with petrol ether, dis- 

 solved in ether, precipitated with petrol ether, filtered and dried in a desiccator, 



Properties of Chlorophyll a. 



Chlorophyll a consists of microcrystalline bunches of lancet-shaped leaflets 

 of a blue- black colour; on powdering they have a green tint. It melts at 

 117-120. 



It dissolves readily in absolute alcohol, ether, benzene, acetone, chloroform 

 and carbon disulphide. The alcoholic solution is blue-green with a deep-red 

 fluorescence; the ethereal solution is blue, greenish when dilute. The 

 solution in carbon disulphide has a yellow tint. 



It is very slightly soluble in cold methyl alcohol, still less in 80 per cent, 

 alcohol and 90 per cent, methyl alcohol. It is soluble with difficulty in petrol 

 ether, but dissolves if alcohol be present. 



A colloidal solution, which has a blue-green fluorescence, is obtained on 

 pouring an acetone or alcoholic solution into water. 



The ethereal solution is decomposed rapidly by shaking with 20 per cent, 

 hydrochloric acid ; with excess of acid the colour is blue. In performing the 

 phase test the colour changes to yellow. 



Properties of Chlorophyll b. 



Chlorophyll b is a microcrystalline solid and forms a powder of a dark- 

 green to green-black colour. It sinters at 86-92 and melts at 120-130. 



It dissolves readily in absolute alcohol, ether and benzene, like the a 

 compom-nt but not quite so easily. The alcoholic and ethereal solutions 

 are green, yellowish compared with those of chlorophyll a. The carbon 

 disulphide solution is yellow-green. The benzene solution in sunlight be- 

 comes red before it is bleached. 



It dissolves with difficulty in cold methyl alcohol, 90 per cent, ethyl 

 alcohol and methyl alcohol. 



It is not so easily decomposed bv hydrochloric acid as chlorophyll a and 



