46 THE BIOCHEMISTRY OF B VITAMINS 



Microbiological tests, in general, rate next to the chemical or physical- 

 chemical tests with regard to speed and accuracy, and have the tre- 

 mendous advantage with respect to natural extracts of very great sensi- 

 tivity and often high specificity. Interference by extraneous substances 

 is always a possibility in every type of assay, and microbiological tests, 

 like all others, are more readily applied to concentrates than to trace- 

 containing mixtures. However, in many cases there is no substantial 

 difficulty in this regard, and extremely small amounts can be determined 

 microbiologically. 



The importance of microbiological tests in vitamin research may be 

 gauged by the fact that by their use pantothenic acid, biotin, pyridoxal, 

 pyridoxamine and folic acid were discovered. They also formed the basis 

 for the discovery of the vitamin properties of niacin, inositol and p-amino- 

 benzoic acid, and for the isolation of the anti-pernicious anemia principle. 



Biological assays using mammals or fowls constituted the first recog- 

 nized vitamin tests, and because of the pre-eminent nutritional function 

 of vitamins, these tests will always remain fundamentally important. 

 Chemical tests and microbiological tests demand that the vitamin to be 

 tested shall be in solution. Animal assays do not have this limitation; 

 and in the sense that vitamins are concerned, by definition, with animal 

 nutrition, the animal tests are the most direct vitamin tests. 



Actually, in animal assays the important factor of availability, as well 

 as presence, comes in. 1 For example, if one were to determine by animal 

 assay the amount of thiamine in raw bakers' yeast, the relatively low 

 result would reflect the unavailability of the vitamin present. From the 

 standpoint of the practical nutrition of animals or human beings the 

 amounts of the various vitamins present in different foodstuffs are of no 

 consequence if the vitamins are unavailable, and only a direct test with 

 animals will give the fundamental information as to how much effective 

 vitamin is present. 



Unfortunately, it is not always safe to carry results obtained using 

 one species of animals over to other species, because the physiological 

 availability may not be the same. If one's interest is in human nutrition, 

 it is necessary to test the materials in question upon human subjects to 

 gain completely trustworthy information. All nutritional tests on animals 

 and humans are complicated by the problem of intestinal synthesis of 

 vitamins, as will be discussed in a later section. 



Biological assays for vitamins using experimental animals are costly, 

 time-consuming and lack accuracy. Nevertheless, for some purposes they 

 are most necessary because vitamins are fundamentally concerned with 

 animal and human nutrition. 



