So much interest is shown nowadays in the relative values of different foods as sources of the different vitamins, that students will welcome a brief account of the procedures and their effectiveness as means of determining quantitatively the factor fat-soluble A.

It would seem logical to test the proposition that young animals, restricted to diets, otherwise satisfactory but containing less than the minimum amount of fat-soluble A necessary for the promotion of growth at the optimum rate, would grow at rates proportional to the supply of this dietary essential, the limiting factor. If this were true, it should be possible on a carefully planned diet to judge from the rate of growth in a number of individuals the nutritive value of that diet with respect to this substance. This possibility has been thoroughly tested by Mc-Collum and Simmonds but the results are disappointing. With suboptimal amounts of fat-soluble A, young rats fail to respond in a manner which can be safely used as a basis of accurate comparison of the amounts tested. Nevertheless, rough approximations of the comparative values of different foods for this dietary factor are possible. Osborne and Mendel (4) and Steenbock and his co-workers (5) have reported studies of this nature which show the great superiority in their content of fat-soluble A of several leafy vegetables over the cereal grains.

A test which has been much used by McCollum and Simmonds (27) is carried out by employing as test animals young rats of the same stock, age and previous nutritive history. They are placed upon a diet which is lacking or essentially lacking in fat-soluble A. A mixture of rolled oats 40.0, gelatin 10.0, a salt mixture 3.7, and dextrin 46.3 per cent, respectively, serves this purpose well. When the initial weights of the animals are about 55 grams, and their ages about 40 days, they usually develop a circular area of puffiness surrounding the eyes in about 30 to 35 days. This sign is premonitory of the onset of ophthalmia, which will now develop within a few days and which is accompanied by excessive secretion in the eyes, and marked edema of the lids. These external signs of ophthalmia are all that need be considered here. Unless the eyes are profoundly damaged recovery takes place on the inclusion in the diet of a suitable amount of fat-soluble A. For the purpose of standardizing conditions with a view to perfecting a quantitative test for this dietary factor they selected, for the addition of the source of fat-soluble A, the time when the puffy ring was easily seen, but when no decided increase in secretion had as yet begun. By comparing the amounts of different food substances given heat or any other treatment tending to destroy the anti-ophthalmic factor necessary to interrupt the course of the disease or to prevent the development of a severe form of ophthalmia, they were able to make fairly satisfactory comparisons of different foods with respect to their curative properties. This method has the advantage of requiring less time and less material than are necessary in a growth experiment. Moreover, the method, they believe, yields results which are more satisfactory. In this method, the animals are brought to the verge of an acute eye disorder, which is averted by an exhibition of a certain quantity of the substance of which their bodies are depleted. The period of observation is shortened to about 6 or 8 weeks.

This method has, however, one element of weakness. A subjective factor is involved in determining the time for beginning the administration of curative measures, but this is reduced to a minimum because of the fairly definite and easily observable incipient edema, and the certainty that once this is seen, the animals will develop severe ophthalmia within a few days.

When small rats are placed on a diet containing traces of the factor fat-soluble A, there is frequently observed a remarkable variation in the time necessary to bring about ophthalmia. In our experience animals weighing 35 grams suffered from it within 3 to 4 weeks, but in one group of 13, individuals were recorded as showing it after 21, 26, 28, 37, 39 days, respectively, while one showed no signs until the 125th day. Since we have no chemical reaction by means of which fat-soluble A can be detected or estimated, the biological test described appears to be the most satisfactory one available.