38 CARBON ASSIMILATION [ch. 



the sol, if concentrated, and then acts, like other weak acids, on the 

 chlorophyll, producing phaeophytin. If the solution is kept neutral by 

 addition of sodium bicarbonate, there is no colour change. The identity 

 of phaeophytin was shown by the spectrum and by the restoration of 

 colour on adding a trace of copper acetate. 



When exposed to light, and the atmosphere in the sealed tube is 

 replaced by oxygen or air, the chlorophyll turns yellow- or brown-green 

 as before and then bleaches. The change of colour from green to yellow 

 or brown is again due to the formation of phaeophytin, this being brought 

 about by the presence of an acid substance, which is produced tluring 

 bleaching, and increases throughout the process. Formaldehyde can be 

 detected in a very slight amount during bleaching, but is formed in 

 much greater quantity after bleaching is complete. 



It is suggested that the formaldehyde is produced by the oxidation 

 and breaking down of the phytol component of the chlorophyll : 

 CH3— CH— CH— CH— CH— CH— CH— CH— C = C— CH.OH 

 ■ I i I I I I I I I 



CH3 CH3 CHq CH3 CH3 CH3 CH3 CH3 CH3 



There is no reason for ascribing to any of the above reactions any 

 part in carbon assimilation. There is at present no hypothesis, supported 

 by satisfactory evidence, as to the process of carbon assimilation. 



Expt. 37. Detection of formaldehyde as a product of oxidation of chlorophyll. 

 Extract 2 gms. of dried nettle leaf powder with 20 c.c. of 80 "/q acetone and transfer 

 it to petrol ether as in Expt. 18. Then shake the petrol ether extract four or five 

 times with an equal volume of 80 % acetone to remove colourless impurities. Next 

 the petrol ether extract is similarly shaken up with 80 "/o methyl alcohol which 

 removes the xanthophyll. This should be repeated until the methyl alcohol is 

 colourless. The petrol ether is finally washed I'epeatedly with water to remove traces 

 of acetone and methyl alcohol. The chlorophyll is in time precipitated as a fine 

 suspension, l)eing insoluble in pure petrol ether. This suspension is filtered through 

 either kieselguhr or powdered talc on a small porcelain filter. The chloi'ophyll is 

 extracted from the powder on the filter with as small a quantity as jMissible of 

 absolute alcohol. This alcoholic solution is then poured, with constant stirring, into 

 100 c.c. of distilled water by wliich means a colloidal solution of chlorophvll is 

 obtained. 



The test to l)e employed for formaldehyde is as follows (Schi-y ver, 4). To 10 c.c. of 

 the liquid to be tested add 2 c.c. of a 1 "/q solution (freshly made) of phenylhydrazine 

 hydrochloride, 1 c.c. of a 5 % solution (freshly made; of potassiinn ferricyanide and 

 5 c.c. of concentrated hydrochloric acid. If formaldehyde is present a pink to 

 magenta colour is develoijed, either deep or pale, according to the quantity of form- 

 aldehyde. The reaction is due to the formation of a condensation product of 

 formaldehyde and phenylhydrazine, and this compound, on oxidation, yields a weak 

 base forming a coloured salt with concentrated hydrochloric acid. The salt is 

 readily dissociated again on dilution of the solution. 



