392 CARNEGIE INSTITUTION OF WASHINGTON. 



vitamine preparation, to the production of which considerable atten- 

 tion has been given during the past two years. 



For many years the proportion of protein which the cells of the green 

 leaves contain has been determined by conventional methods and, ap- 

 parently, in no case has any corresponding quantity of protein been 

 isolated and chemically identified. In consequence there is at present 

 no satisfactory evidence of the nature of the compounds containing a 

 large proportion of the nitrogen in the green leaves of any plant. The 

 highly specialized physiological functions of the leaf justify the expec- 

 tation that it may contain constituents chemically unUke those found 

 in other parts of the plant or in the cells of animal tissues. How far 

 such constituents belong to types of compounds peculiar to the leaf, or 

 to types already well known, is scarcely known at present. It is 

 highly probable that the leaf is the seat of protein synthesis, as already 

 it is known to be the seat of carbohydrate synthesis. The protein of 

 the leaf may represent the original protein from which all other kinds 

 are formed, either du-ectly or indirectly. 



Apart from numerous attempts to apply conventional methods for 

 distinguishing non-protein nitrogen from protein nitrogen in leaves and 

 green plants, little appears to have been done to increase our knowledge 

 of this subject. Botanists have made microscopic studies of leaves 

 and other parts of plants which furnish some facts of interest respecting 

 their protein constituents, but at present these reveal little with regard 

 to their chemistiy or value in nutrition. The experience which we have 

 had in the past in studying the proteins of seeds prompted us to make 

 this investigation, which finally showed how most of the proteins can 

 be easily separated from the fresh leaf. Although preparations thus 

 obtained doubtless are mixtures of several individual proteins, they are 

 more suitable for a study of the nutritive value of the total protein 

 furnished by the leaf than M^ould be any single protein contained in 

 this mixture. 



Spinach leaves were employed because these contain more nitrogen 

 than do most other leaves, and also because a fresh supply can be ob- 

 tained throughout the greater part of the year. ^Vhen the walls of the 

 cell are broken by grinding, its contents are liberated and a mixture 

 of the nuclei, chloroplasts, cytoplasm, sap, debris of cell-walls, etc., is 

 obtained. The parts contributing to this mixture are no longer in the 

 same relation to one another as when intact within the living cell; 

 they form an amorphous mixture, in which the components no longer 

 can be recognized under the microscope. When this mixture is diluted 

 with one or two volumes of water and either centrifuged at high speed 

 or passed through coarse filter-paper, an opaque green, slightly viscid 

 fluid is obtained and also a residue, consisting chiefly of the cell-walls. 

 Although the fluid part is very opaque and looks as if it contained much 

 suspended solids, microscopic examination reveals nothing except ex- 



