124 Scientific Proceedings (119). 



cartilages contained deposits of calcium salts. We have finally 

 worked out a diet which fulfills the requirements for the test. 1 



Diet (Lot) 3143. 



Wheat 33.0 



Maize 33.0 



Gelatin 15.0 



Wheat gluten 15.0 



NaCl 1.0 



CaCOs 3.0 



This diet is faulty because it contains only 0.3019 gram of 

 phosphorus in 100 grams of the mixture, while calcium is present 

 to the extent of 1.221 grams per 100 grams of food. We have 

 found that about 0.40 gram of phosphorus per 100 grams of food 

 represents the lowest concentration of this element on which 

 optimal nutrition can be secured when 0.641 gram of calcium 

 are present in each 100 grams of ration. The fat-soluble A is 

 low in this diet (Lot 3143), but it is sufficient to allow growth and 

 to protect the rat against xerophthalmia. The diet also contains 

 very little of an uncharacterized organic substance present in 

 certain fats. We have previously discussed in detail the results 

 of restricting rats to it. 



Technic of Test. 



A group of young rats is placed on diet 3143 for 35-45 days, 

 or until they begin to lose control of their hind legs. They are 

 then divided into two groups, a control group which continues to 

 receive diet 3143 unchanged, and a test group, which is given 

 for the number of days deemed necessary the faulty diet plus the 

 substance which is to be tested. 



When a sufficient number of days have elapsed for the test 

 substance to have produced its effects, the animals are killed and 

 the bones which are to be studied are split longitudinally. The 

 proximal end of the tibia is best for the purpose. One half of 

 the bone is immersed in a dilute silver nitrate solution and exposed 

 to light. It is then examined in the solution through a binocular 

 microscope for the presence of a newly formed line of calcification 

 in the proliferative cartilage. This line of calcium, which looks 

 like the cross section of a honeycomb under strong magnification, 

 is blackened by exposure to light. If this line is present, the test 



1 Shipley. P. G., Park, A. E.. McCollum, E. V.. and Simmonds, Nina. Proc. 

 Soc. Exper. Biol, and Med., 1921, xviii, 277. 



