Prevention of Rickets in the Rat. 



123 



60 (1807) 



A delicate biological test for calcium-depositing substances. 



By E. V. McCOLLUM, NINA SIMMONDS, P. G. SHIPLEY, and E. A. PARK. 



[From the Department of Chemical Hygiene, School of Hygiene and 

 Public Health, and from the Department of Pediatrics, 

 Johns Hopkins University, Baltimore, Md.] 



Some time ago we described two diets, Nos. 2638 and 2677, 

 which we used with a view to the development of a biological test 

 which would show the calcium-depositing power of any given 

 substance. 1 The test was carried out as follows: The faulty diet 

 was first fed to a group of young rats for the purpose of making 

 the epiphyseal cartilage free from calcium, and producing a 

 rachitic metaphysis. After a sufficiently long period had elapsed, 

 the test substance was added to the diet of those animals which 

 were to serve as test subjects, while the faulty diet without the 

 test substance was continued in the case of the control rats. 

 Substances, which when added to the faulty diets enabled the 

 organism to deposit lime salts, caused the reappearance of the 

 provisional zone of calcification in the bones. This biological test 

 we called the "line test," because the new provisional zone of 

 calcification appeared as a line of calcium salts extending trans- 

 versely across the bone with a limeless cartilage on one side of it 

 and a limeless metaphysis on the other. 



The success of this test depends on the use of a diet which 

 uniformly causes the epiphyseal cartilage and the metaphysis to 

 be free from calcium salts. It is not sufficient that a diet should 

 merely produce rickets. The rickets which it produces must be 

 of so severe a type that no vestige of calcium remains in the 

 cartilage, and a wide metaphysis is formed. Moreover, the diet 

 must be so constituted that the animals restricted to it will grow 

 and maintain a fair state of general health and nutrition. The 

 diets which we earlier described were not satisfactory, since they 

 did not invariably produce typical rickets. Now and again an 

 animal which was restricted to one of them would be found whose 



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

 H. T. Jour. Biol. Chem., 1921, xlv, 343. 



