Aug. 1, 1925 
Vitamin A in Beef, Pork, and Lamb 
205 
In the earlier work reported in this paper, both the starch and the 
yeast were thoroughly extracted with ether before being used, but 
this practice was discontinued as soon as it was found to be unneces¬ 
sary. Both the crude casein and the dried ox muscle used as sources 
of protein were found to contain considerable vitamin A and purifi¬ 
cation was required. Purified ox muscle was used in most of the 
tests reported in this paper, but later it was found more satisfactory 
to purify casein, and that product was then used exclusively as the 
supplementary source of protein in these vitamin A studies. 
The ox muscle was purified as follows: Water-insoluble muscle tis¬ 
sue, a by-product resulting from the preparation of beef broth for 
the growth of tubercle bacilli, was dried, ground fine, and extracted 
with ether in large percolators until the material was practically 
free from fat. Tiie extracted muscle was mixed with 60 per cent 
alcohol by weight in a large dish, transferred to large percolators, 
and extracted with 60 per cent alcohol until the extract was of a very 
light straw color. It was found to be practically impossible to obtain 
a colorless extract. The extraction with ether required two or three 
days and that with alcohol four or five days. The extracted material 
was dried and tested for vitamin A, but was found still to contain an 
appreciable amount of the vitamin. In order to reduce the vitamin 
content still further, the extracted muscle tissue was then heated in a 
current of air at an average temperature of approximately 115° C. 
for 24 hours, when it was found to be practically free from vitamin 
A. This method was found to be very tedious and later was 
abandoned. 
The following method has been used regularly since October, 1923, 
in the purification of commercial casein, with very satisfactory 
results. Six hundred grams of granular casein is spread out in a thin 
layer in a shallow, galvanized-iron pan 183^ by 25 inches by 3^ inch 
deep, and five of these pans are set at one time in the drying oven 
shown in Plate 1, D ana E. The casein is then heated in a current 
of air at an average temperature of 115° C. for 24 hours. The pans 
are then taken out and the casein is transferred to a large dish, thor¬ 
oughly mixed, and returned to the pans. Heating is continued for 
another 24 hours, when the casein is nearly always found to be free 
from vitamin A as determined by feeding tests with rats. If the 
rats make more growth than is considered normal for the rats on a 
vitamin-A-free ration, the casein is heated for an additional 24 hours 
and tested again. Ordinarily about 25 pounds of casein is purified, 
thoroughly mixed, and tested in advance of requirements for the 
product. The heating changes the color of the casein to a very light 
brown but does not char the product. Comparative feeding tests 
have shown that rats fed a ration containing 20 per cent of protein 
from the purified casein made equally as good growth as those tha£ 
received a ration containing a like percentage of the untreated casein, 
the rations being adequate in other respects. 
Feeding Tests with Basal Rations 
Five lots of purified ox muscle and one lot of purified casein, 
which comprise all of these products that were used as sources of 
protein in the feeding tests reported in this paper, were tested to 
determine their freedom from vitamin A, with the results shown in 
