VITAMIN A 227 



observations which apparently yield discordant data as to whether vita- 

 min A is soluble in ether, and as to whether it occurs in nature in 

 different forms having different solubilities. As yet, however, there seem 

 to be differences among the recorded observations which are only in 

 part explainable on this basis alone. Thus the observations of Osborne 

 and Mendel just mentioned would seem to call for a revision or reinter- 

 pretation of the extraction experiments of McCoUum, Simmonds and 

 Pitz (1916b) mentioned above, and are also difficult of coordination 

 with observations subsequently recorded by Steenbock and Boutwell 

 (1920b) who attempted to concentrate the fat-soluble vitamin* from 

 carrots, alfalfa, and yellow corn by fat solvents with the following 

 results : 



The fat-soluble vitamin in carrots was not removed to any extent 

 by saturation with corn oil and extraction with ether, and apparently 

 not at all by similar saturation with lard. The vitamin appeared to be 

 slightly soluble in ether, somewhat soluble in chloroform and carbon 

 disulfide, and quite soluble in alcohol and benzene. With alfalfa, more 

 satisfactory results were obtained with ether, benzene, and alcohol, 

 sufficient fat-soluble vitamin being obtained in the ether extract to 

 permit satisfactory growth when fed to rats at a level corresponding to 

 20 per cent of alfalfa in the food mixture. These authors also employed 

 with the alcohol extract from alfalfa meal the method commonly used 

 for the separation of carotinoids. The extract was saponified with 

 alcoholic potassium hydroxide and the mixture extracted repeatedly with 

 ether until the yellow pigments were completely removed. The ether 

 extracts, after washing with water to remove alkali and salts, were 

 evaporated on a water bath to a small volume. This extract proved to 

 be rich in fat-soluble vitamin. A further concentration of the vitamin 

 was brought about by taking up a similar extract in a mixture of alcohol 

 and petroleum ether and subjecting it to fractional extraction. The 

 petroleum-ether-soluble carotene fraction was found to contain an 

 abundance of the vitamin, while the alcohol-soluble xanthophyll fraction 

 contained little or none of it. 



About the same time Zilva (1920) reported experiments in which 

 fractions active in vitamin A were obtained from fresh vegetables 

 (carrots and cabbage) by extraction with alcohol. Absolute alcohol ex- 

 tracted in the form of a sirup about 7.5 per cent of dry matter from 



* Inasmuch as Steenbock found subsequently that under the conditions of testing used in 

 his laboratory there was danger of confusion between vitamins A and D, it is conceivable 

 that vitamin D may have been the growth-limiting factor in the experiments of Steenbock and 

 Boutwell, whereas Osborne and Mendel were dealing with vitamin A. Investigations upon the 

 quantitative differentiation of vitamins A and D are discussed further on in this chapter and 

 in the chapter which follows. 



