136 OCCURRENCE OF FREE AMINO ACIDS — INSECTS 
AMINO ACIDS AND DERIVATIVES IN DROSOPHILA 
HERSCHEL K. MITCHELL anv JOHN R. SIMMONS 
California Institute of Technology, Division of Biology, Pasadena, Calif. (U.S.A.) 
A number of years ago HADORN AND MITCHELL! presented data showing changes in 
patterns of amino acids and related substances that occur during development in 
Drosophila melanogaster. Two prominent components among the amino acid deriva- 
tives were shown to be peptides containing several amino acids and in subsequent 
work by HAporN, CHEN et al.2~* at least three more peptides were observed. In addi- 
tion, Fox® reported a peptide difference in male and female Drosophila, and evidence 
for the existence of peptides among the lipid~amino acid conjugates of the fly was 
also obtained’. 
It is of importance to point out that nearly all of the observations mentioned 
above were made by the use of paper chromatography, and, although the method has 
yielded a wealth of information, it has limitations in resolving power and, at concen- 
trations below the level of overloading the paper, it reveals only the most abundant 
components to be found in extracts. Consequently when we undertook the work to 
be summarized here we resorted to ion-exchange chromatography followed by paper 
chromatography, electrophoresis and other methods suitable for purification of 
natural substances. Our objectives were, first, to examine in greater detail the pattern 
of ninhydrin-reacting substances in Drosophila including components present in low 
concentrations; second, to determine the structures of peptides already observed as 
well as of others that might be found; and third, to study the metabolism of the 
peptides particularly in relation to a possible role in protein synthesis. That all these 
objectives have not been achieved will become evident but some 5 years of exploration 
has nevertheless, yielded much useful information. 
PRELIMINARY OBSERVATIONS 
In a study of the occurrence of peptides as normal constituents of tissues it is essential 
that, during extraction, there is no possibility of significant hydrolysis of protein or 
existing peptides. Some idea of the magnitude of this problem is illustrated in Fig. r. 
As shown, incubation of a larval homogenate even at 0° yields an appreciable increase 
in ninhydrin-reacting material while a very large increase occurs in a few minutes 
at 28°. We have therefore made use of extraction at —20° as the first step. It 
has also been observed that, under some conditions, extraction by heating causes 
loss of ninhydrin-reacting material presumably due to the presence of reactive amino 
acid derivatives. We have therefore avoided heating as much as possible. In initial 
experiments use was made of Dowex-50-4X and a modification of the gradient elution 
system of THompson’. Results from one such experiment are summarized in Fig. 2. This 
is representative from seven separate runs in which total propanol-water (1 : 1, —20°) 
References p. 146 
