PIGMENTS IN THE CHLOROPLASTS 383 



pigments. Probably they are kept floating by the hydrophilic properties 

 of the proteins. Dihition with organic solvents breaks the pigment- 

 protein link, denatures and precipitates the proteins, and dissolves the 

 chlorophyll and the carotenoids. 



Pure organic solvents — e. g., ether, water-free acetone, or alcohol — 

 do not dissolve chlorophyll from the leaves. One could suggest that the 

 solvents are unable to break the pigment-protein link so long as no 

 water is present to take care of the proteins. However, the immersion 

 of leaves into ether causes a shift in the position of the absorption bands 

 and an increase in fluorescence, which seems to indicate that the pigment 

 has been liberated from the protein complex and has passed into a lipoid 

 phase. What forces still prevent it from diffusing from the cells into 

 ether is not immediately clear. 



The only efficient way to extract chlorophyll and other pigments from 

 the cells is by using aqueous organic solvents, water disintegrating the 

 proteinaceous fraction of the chloroplast structure, and the organic sol- 

 vent taking care of the lipoid fraction, including the pigments. Once 

 separated from the cell structure, the pigments become easily soluble in 

 pure organic solvents. 



Several other observations speak for the association of chlorophyll 

 with some "carrier" in the cell. One is the position of its absorption 

 bands, which are shifted 10-20 mju towards longer waves relative to 

 their position in solution (cf. Vol. II, Chapter 22); e. g., the main absorp- 

 tion peak in the red is situated at 675-680 m/x in the living cells, as 

 against 660-670 mn in organic solvents. The fluorescence band also is 

 shifted towards the red (cf. Vol. II, Chapter 24), and the fluorescence is 

 at least ten times weaker than that of dissolved chlorophyll. The ab- 

 sorption bands of the carotenoids are shifted even more strongly than 

 those of chlorophyll (cf., for example, Menke 1940). 



Chlorophyll in the living cell is much less sensitive to acids than is 

 chlorophyll in solution. (According to Hilpert, Hofmeier, and Wolner 

 1931, it is more sensitive to cold dilute alkali.) It is also much more 

 resistant to bleaching (photoxidation or photoreduction; cf. Chapter 19, 

 page 537). This stability, too, points to the protective action of a 

 "carrier," probably a protein. Inman and Crowell (1939) found that 

 trypsin causes the conversion of chlorophyll in the cells into pheophytin, 

 and suggested that magnesium serves as a link between chlorophyll and 

 protein. Zirkle (1926) found that proteins in etiolated chloroplasts are 

 easily digested by enzymes, while those in green chloroplasts are more 

 resistant; thus while chlorophyll is protected chemically by the proteins, 

 the proteins are protected by chlorophyll. 



When leaves are put into ether, or cooled by liquid air, or boiled in 

 water, the absorption band is shifted toward its position in true solution. 



