704 
Miller.—A Physiological Study of the 
cells of the cotyledons were now apparently free from all reserve material. 
They had enlarged and had the appearance of the parenchyma cells of an 
ordinary vegetative leaf. At this stage the plant has evidently become 
wholly independent. 
Stage V. The age of the seedlings used at this stage was thirteen 
days, and during the last seven of 
these the plants were kept in the C 0 2 - 
free apparatus. The seedlings at this 
stage differ very little from those of 
the previous period. The plumule 
had lengthened and the first internode 
had a length of o-6 to i-2 cm. The 
hypocotyl had lengthened a little and 
more copious side roots had developed. 
The seedlings for several days previous 
to the end of this period had shown 
no further growth of their parts. This 
was to be expected, since the plants, 
being cut off from the supply of 
nourishment both from the soil and 
the air, had no material for growth 
after the reserved material had been 
consumed and were slowly starving. 
See Fig. 6. 
ANALYTICAL METHODS. 
To avoid the oxidation of the 
oil during the preparation of the 
material for analysis, the following 
modification of Lowensteins ( 31 ) 
method was used. After the wet 
material had been thoroughly ground 
and pulverized in a mortar, it was 
transferred to broad, shallow pans on 
the water-bath and treated with twice 
its volume of 95 % alcohol. After the 
alcohol had evaporated the process 
was repeated a second and a third 
time and again with absolute alcohol. 
After the evaporation of the alcohol the material was placed in the 
oven at ioo°-io5 ° C. for 20 to 30 minutes and then transferred to the 
desiccator and finally to tightly closed vessels until needed for analysis. 
Fig. 6 . Stage V of Seedlings. 
