260 THE VITAMINS 



Experimental Procedure. — The discussion which follows relates to 

 young albino rats which, unless otherwise stated, were from families 

 fed upon a diet of one-third dried whole milk and two-thirds ground 

 whole wheat, with sodium chloride in the proportion of 2 per cent of 

 the weight of the wheat, with or without the addition of 30 to 60 grams 

 per week of fresh lean beef per adult. This addition of lean beef was 

 found to have no appreciable effect upon the amount of vitamin A 

 stored in the body of the young rat as judged from the length of time 

 it was able to survive when placed upon vitamin-A-free diet. 



When such 21- to 29-day old rats of an initial weight of 35 to 50 

 grams are placed upon a diet well adapted to their needs in other 

 respects but devoid of vitamin A, such as our basal vitamin-A-free diet 

 described earlier, growth usually continues for from four to five weeks 

 during which period our rats usually gain from 35 to 50 grams. Around 

 the time when cessation of growth is anticipated the animals should be 

 weighed every other day and finally daily. Having decided upon a 

 definite and uniform diet for the families from which the young are 

 to be drawn, and a uniform age at which to place the young upon the 

 vitamin-A-free diet, one can learn by experience about when to expect 

 cessation of growth, and will also be guided by the general appearance 

 of the animals in determining when the fore-period should end and 

 the test period begin. This should be after the animals have certainly 

 used up their previous reserve store of vitamin A but before they 

 have been permanently injured by the vitamin- A deficiency. 



As soon as the body weight ceases to increase, and/or the first signs 

 of eye disease appear (slight swelling of the eyelids and an accumula- 

 tion of exudate in the corners), usually accompanied by a barely per- 

 ceptible tendency to flabby musculature and slightly unkempt fur, the 

 depletion period should be ended. 



Individuals or litters which appear too weak or too vigorous to be 

 comparable to the general average should be discarded. The others are 

 placed in individual cages with raised screen bottoms to prevent access 

 to excreta, and at least one of each litter is continued on the basal 

 vitamin-A-free diet until death, as a "negative control," while the others 

 are fed graded portions of the food to be tested, as their sole source 

 of vitamin A, daily or at other suitable intervals during the test period 

 in addition to the basal diet and distilled water ad libitum. Any advan- 

 tage in survival period or maintenance or gain in weight shown by a 

 test animal over its "negative control" must be attributed to the vitamin 

 A received from the weighed portions of the food or other material 

 which is being tested, provided the experiments are conducted properly 



