I44 H. K. MITCHELL AND J. R. SIMMONS 
IDENTIFICATION OF PEPTIDES 
At the present time we have done relatively little work to determine the composition 
of the peptides observed and no work on amino acid sequences. There are too many 
and they are much alike, making the isolation of pure substances a large and tedious 
program. This is illustrated by the information in Fig. 7 which we present, with 
some misgivings, since we are aware of the limitations and inaccuracies of the methods 
lyr Asp 
80'000 < <4 
60000 
40b00 
20000 
MILLIMICROMOLES 

FRACTION 
Fig. 8. Amino acids and peptides in rabbit brain. Shaded areas represent quantities before hydrolysis 
and black areas the increase on hydrolysis. The data are plotted at three levels with 20-fold differ- 
ences in concentration to show the presence of numerous peptides at relatively low concentrations. 
Extract: Fraction 1 prepared as in Fig. 3. Column: 1.6 x 16cm, Dowex-1-8X treated with 4 M 
sodium acetate and washed thoroughly with water. Elution: water to tube No. 5 (1o-ml fractions) 
followed by a gradient produced by introducing 0.5 MW triethylamine acetate (pH 5.0) into roo ml 
of water in a mixing vessel. This system is designed for separation of acidic compounds and elution 
positions of some known substances are shown in the graph. 
used. Samples were taken of acidic components from a column separation (as in Fig. 2) 
and components were purified further by chromatography and electrophoresis on 
washed paper. After homogeneity seemed assured samples were hydrolyzed and 
analyzed by one- and two-dimensional chromatography and by photometric deter- 
minations of quantities. Greatest uncertainties are in the ratios when one amino acid 
is repeated several times and, of course, in original purity. 
These results emphasize the separation problem since many of the 42 components 
listed are very much alike, some series differing only in the number of repeats of 
glutamic acid. The predominance of non-essential amino acids is also obvious. 
References p. 146 
