INCORPORATION OF "C INTO TISSUE 807 



lesterol carbon into barium carbonate would have resulted in a still greater reduc- 

 tion of the activity figures. 



The fat-free tissue, after being repeatedly washed with water was repeatedly 

 treated at 90° C for 10 min. with 5% trichloracetic acid to obtain the raw proteins. 

 These were abundantly rinsed with water, then dried by washing with alcohol 

 and ether. Subsequently the activity of the protein samples was compaied. 



We purified in our later experiments the total fat samples with colloidal iron^^\ 

 In the case of liver fats such a purification leads to an increase in the activity 

 figures. In experiments taking 3 hours, an average increase of 11 per cent was 

 observed, while Dr. Beeckmans working in this laboratory found in experiments 

 taking 15 minutes a 7 per cent average increase. Treatment with colloidal iron 

 removes thus less active constituents of the liver fats, possibly urea, amino 

 acids and other impurities. Some phosphatides are removed by their treatment 

 as well. In experiments taking not more than a few hours, the activity of phospha- 

 tides is less than the activity of the total fats of the same weight, the ratio of the 

 activity of 1 mgm total fat and 1 mgm phosphatide being about 1.3. Some removal 

 of less active phosphatides in the course of the purification process with colloidal 

 iron will thus also lead to an increase in the activity of the total fats of the liver. 



Purification with iron has very different effects on brain fats. After purification 

 their activity decreases with 11 per cent in experiments taking 3 hours. More active 

 fractions are thus removed by this process. The decrease in the activity figures 

 due to purification is much larger in experiments of short duration. In experiments 

 taking 15 min. only, treatment with colloidal iron removes 3/4 of the ^*C content 

 of the total fats, 45 % of that of the muscle fat and 1/3 of the intestinal mucosa. 



The dry Uver weight of fed mice weighing 22.7 gm (mean value of a few hundred 

 animals) was 0.31 gm. Liver dry weight amounted to 28.2 % of its fresh weight. 

 We found the fat content of dry Uver tissue = 21.7%, that of fresh tissue = 7.4%.^^^ 

 In the Uver of fed 15 gm mice Stetten and Grajl^^^ found a fatty acid content of 

 4.5%. From the above data it follows that 61% of the liver fats is made out of 

 fatty acids. 17.1% of the weight of the fresh Uver was made out of proteins. 



When comparing the activity of the dry total tissue or protein samples secured 

 from irradiated mice and from controls, 50 mgm of each to the dried samples we 

 placed in an aluminium dish of 1.2 cm diameter and the activity of the samples 

 was compared. The small size counters used had a mica window of about 2 mgm 

 per cm^ and a background of about 3 counts per minute. 



The fat samples, the activity of which was appreciably larger than that of the 

 total dry tissue or protein samples weie placed in Ottesen's micro dishes, which 

 proved to be very suitable for this purpose. The counts measured for an infinitely 

 thick fat sample placed in a dish of 1.2 cm diameter made out 4.8 times the number 

 of counts measured when the infinitely thick fat sample was placed in a micro 

 dish. 



In some cases we burnt our samples and compared their activity after converting 

 their carbon into BaCOj. In view of the work involved in this procedure (we 

 measured the activity of a very great number of samples) and also because the 

 low carbon content of BaCOg we preferred to compare the activity of our samples 

 without converting them into barium carbonate. Total Uver fat samples, for 

 example, which were found to contain 65.2 per cent carbon, while the carbon 



^i) J. FoLCH and D. Van Slyke, Proc. Exp. Biol. Med. 41, 514 (1939). 

 ^2) H. C. Hodge, P. L. MacLachan, W. R. Bloor, E. Welch, S. L. Kornberg 

 and M. Falkenheim state a fat content of 7%. J. Biol. Chem. 67, 137 (1948). 

 (3) D. Stetten Jr. and G. F. Grahl, J. Biol. Chem. 148, 509 (1943). 



