478 
Journal o f Agricultural Research 
Vol. XXIX, No. 10 
The most plausible explanation of 
the decline in acidity seems to be that 
this is due to neutralization of the 
originally free, and later liberated, 
COOH groups by basic groups which 
have not yet been attacked by for¬ 
maldehyde. This explanation depends 
upon the possibility of basic groups 
existing in the presence of formalde¬ 
hyde. That such a possibility exists 
has been shown by Schiff ( 14 ), who 
reported that relatively few NH 2 
groups of proteins are combined with 
formaldehyde, and by Proctor (10 , p. 
14 &), who has shown that the reaction 
between amino-acid anhydrides and for¬ 
maldehyde, and also between histidine 
and formaldehyde, is irregular; that 
is, not all of the NH 2 groups are 
combined. 
If the assumption that the decline 
in acidity of formolized serums is due 
to a neutralization of the COOH 
groups of the protein by NH 2 or other 
basic N groups be accepted, then the 
reason for the applicability of the 
formula 
which applies only when the reacting 
substances are present in equimolecu- 
lar proportions, becomes apparent: pro¬ 
teins are composed of amino acids; and 
practically all amino acids contain basic 
and *acid groups in equimolecular 
quantities. 
THE P H VALUE OF FORMOLIZED 
SERUMS 
The results obtained in the titration 
studies suggest, of course, that the re¬ 
actions which result in the transforma¬ 
tion of the more soluble proteins of 
formolized serums into less soluble 
forms may be caused by changes in 
the hydrogen-ion concentration in¬ 
duced by the presence of added for¬ 
maldehyde. The following two experi¬ 
ments bear on that question. 
EXPERIMENT V 
A hog-blood serum was divided into 
two portions. To one, 0.09 per cent of 
formaldehyde was added in the form 
of a solution of formaldehyde, and to 
the other 0.5 per cent of phenol was 
added. The euglobulins and the P H 
value 7 of the serums were determined 
at the intervals shown in Table VII. 
It will be noted that following the 
addition of formaldehyde and phenol, 
respectively, an initial decrease in the 
P H value took place, and that this was 
in each sample followed by an increase. 
In the formaldehyde-treated fraction 
the euglobulins increased 124 per cent, 
while the phenol fraction increased 
but 24 per cent. This suggests that 
the changes in the solubilities of the 
proteins of formaldehyde-treated se¬ 
rums are not due solely to changes 
in the P H value. 
EXPERIMENT VI 
A horse-blood serum was divided 
into two portions, to one of which 0.74 
per cent and to the other 1.85 per cent 
of formaldehyde were added. The 
P H value and the titrable acidity of 
these serums were determined at the 
intervals and with the results shown 
in Table VIII. 
value and euglob - 
Table VII .—The effect of formaldehyde and phenol on the Ph 
ulin content of hog-blood serum 
Added 
1 Time of contact 
P 11 
Euglobulins 
(grams per 
100 c. c.) 
Nothing.... 
i Fresh 
8.4 
1.41 
1.81 
0.09 per cent formaldehyde_ 
! /1 day.. 
8.0 
j\l month_ 
8.3 
3.16 
0.05 per cent phenol__ 
jjl day_ 
8.0 
1.60 
a 1 mon th_ _ 
8.5 
1.75 
1 1 
7 Determined colorimetrically by Dr. F. W. Tilley, of this division. 
