Comparative Stiidij of Chlorophyll. 413 



that Kraus' band IV. is very evident in spectrum 5, without 

 there being, however, any fusion of the first three bands ; the 

 alcoholic extract represented in spectrum 5 was made from 

 Spirogyra material which had been washed in distilled water, 

 then boiled for ten minutes, washed again in distilled water, 

 and extracted with absolute alcohol. The explanation of the 

 difference between the two just-mentioned extracts I would 

 suggest is that in fresh material the green colouring-matter 

 is a protoplasmic, hence an alkaline body, and that absolute 

 alcohol extracts the colouring-matter so rapidly as to restrict 

 to a minimum the action of post-mortem changes on the 

 alkaline chlorophyll, amongst which changes one of the first 

 seems to be that the alkaline protoplasm becomes acid. In 

 material, on the other hand, which has been boiled, the 

 chlorophyll will be exposed to post-mortem changes, the acid 

 ceU sap will be able to act on the chlorophyll as well as 

 any gases that may be in the water, such as carbon dioxide, 

 &c. ; and the very act of prolonged boiling will bring the 

 material we are treating in contact with the oxygen of the 

 air, and thus the chlorophyll will change its composition, 

 and the appearance of the band in the green will be the 

 consequence. Now, one might ask, does not the boiling 

 itself affect the chlorophyll, cause its decomposition, and the 

 appearance of the band ? This does not seem to be the 

 case, for I boiled an alcoholic extract made from fresh 

 material, according to my method, for eight minutes, but 

 could not detect any change in the band in the green part 

 of the spectrum ; the band did not become darker. What 

 the effect of an alkali is on an extract of chlorophyll, made 

 with absolute alcohol from boiled material, with regard to 

 Kraus' band IV., is readily seen by comparing spectra 5 

 and 6. In spectrum 6 the band is much paler than in spectrum 

 5, for three drops of liquor ammonii were added to a test- 

 tube full of the extract, and the extract then heated up to the 

 boiling-point for one minute. It is evident, therefore, that a 

 strong alkali, such as ammonia, will decrease the amount of 

 absorption in the green part of the spectrum ; an acid, on the 

 other hand, e.g., nitric acid, will increase the intensity of absorp- 

 tion, as shown in spectrum 7 ; but great care must be taken 

 in adding a strong acid, or the whole of the spectrum will be 

 changed, and the band in the green will become paler again. 



