SWEET-CLOVER SEED. _ 83 



MICROCHEMISTRY OF THE SEED COAT. 



Tests for cutin showed that there was very little present in the 

 seed coat. Slight reactions for cutin were observed in the cuticle, in 

 the outer margin of the cuticularized layer, and in the basal portion 

 of the cones. These reactions were so slight as to be almost negli- 

 gible. It is evident that the cuticle and cuticularized layer are not 

 well named in Melilotus alba and M. officinalis. Tests for cellulose 

 showed that it was present in the cuticle, cuticularized layer, cones, 

 the walls of the Malpighian cells below the light line, and the walls of 

 the cells of the osteosclerid and nutrient layers. (PI. V, fig. 9.) 

 The reaction for cellulose in the Malpighian cells was quite distinct in 

 the walls below the light line, less distinct in the cones and cuticle, 

 and least distinct in the cuticularized layer. 



Tests for lignin occasionally showed slight traces in the Malpighian 

 cells below the light line. When treated with reagents for pectic sub- 

 stances, the cuticle, cuticularized layer, cones, and all cell walls 

 below the light line gave a definite reaction. The reaction of the 

 cones and cuticle was more pronounced than the cuticularized layer. 

 Tests for callose gave no reaction except in the upper part of the 

 light line. This part of the light line stained slightly blue with 

 aniline blue and was easily dissolved with sulphuric acid. In cutting 

 free-hand sections of fresh material the Malpighian layer sometimes 

 broke along this line. The greater part of the light line reacted to 

 none of the tests, and its chemical nature was not determined. 



When microtome sections of seeds in different stages of develop- 

 ment were treated with various stains, the results were in accord 

 with those obtained with free-hand sections. Thus with safranin 

 the periphery and cones of the Malpighian cells were slightly stained, 

 while haematoxylin and methyl blue stained all the seed coat except 

 the light line. The cones and cuticle stained more readily than the 

 cuticularized layer, but neither stained as deeply as the cell walls 

 below the light line. Methylene blue, methyl violet B, and mauvein 

 stained all above the light line, indicating the presence of pectic sub- 

 stances; however, the staining was more prominent in the "cones and 

 cuticle. 



The difference in reaction of the cones and cuticularized layer to 

 the cellulose and pectose tests probably indicates a difference in 

 density rather than a difference in chemical composition. Since the 

 cuticularized layer separates readily from the cones, there may be a 

 difference in physical properties. 



With Congo red the upper part of the light line was only very 

 slightly stained, but aniline blue had a more pronounced effect. 



The microchemical tests applied to the seed coat show that in the 

 region above the light line there is only a slight trace of cutin or 



