384 CHLOROPLASTS AND CHROMOPLASTS CHAP. 14 



thus indicating the probable decomposition of the protein-chlorophyll 

 complex (Willstatter and Stoll 1918; Seybold and Egle 1940). Chloro- 

 phyll in leaves killed in this way is much more sensitive to oxygen and 

 acids than it was before killing. 



It thus seems certain that chlorophyll (and the other chloroplast 

 pigments) are associated, in the living cell, with the cell proteins, and 

 probably also with some lipophilic compounds. We now ask: is this an 

 association in stoichiometric proportions; and does it involve uniformly 

 all the chlorophyll contained in the cell? 



That the association of porphine derivatives with proteins can lead 

 to the formation of stoichiometric compounds is well known from the 

 example of hemoglobin and cytochrome, in which one porphyrin molecule 

 is associated with one so-called "Svedberg unit" of protein (molecular 

 weight ~ 17,000). Whether an association with lipides, which is 

 produced by nonpolar, van der Waals' forces, also can result in stoichio- 

 metric relations is less certain. 



As early as 1886, Reinke speculated that chlorophyll (and the yellow 

 pigments) may be bound to proteins in the leaf in the same way as 

 hemin is bound to globin in hemoglobin. Since then, it has become 

 evident that most biological catalysts (enzymes) consist of similar 

 combinations of a protein "carrier" with an active ("prosthetic") 

 molecule. Since chlorophyll may be considered as an enzyme which 

 becomes active in light, the hypothesis that it has a similar "chromo- 

 proteic" constitution appears natural. However, this hypothesis still 

 lacks definite confirmation. Many different chlorophyll-protein suspen- 

 sions and colloidal solutions have been prepared, both by disintegration 

 of plant material and by the interaction of pure pigment with proteins 

 in vitro; but none had a simple and reproducible composition similar to 

 that of hemoglobin or cytochrome. Difficulties have been encountered 

 also in reproducing in chlorophyll-protein complexes the two above- 

 mentioned properties of "natural" chlorophyll^the position of its 

 absorption band and its fluorescence. 



2. Pigment-Protein Suspensions and Solutions 



The preparation of green aqueous extracts from leaves was first 

 described by Herlitzka in 1912. He obtained them by the grinding of 

 spinach leaves, and described them as possessing the unchanged spectral 

 properties of chlorophyll in the leaves. Lubimenko (1921, 1927) found 

 that certain species, e. g., Aspidistra elatior and Funkia, are particularly 

 suitable for extraction by water, and give stable clear "solutions" which 

 are not precipitated by centrifugation. The spectrum of these solutions 

 was "identical with that of the leaf"; they were stable in sunlight; and 

 acetone or alcohol precipitated from them the protein-chlorophyll 



