VITAMINS 365 



sorbed is directly related to the concentration and is used to estimate 

 the latter. Most such measurements do not require much material, 

 and what is used is readily recovered after the analysis. Spectrophoto- 

 metric procedures are widely employed in determining a great variety 

 of materials and are closely related in theory to the colorimetric 

 methods of the previous section. 



Microbiological Methods 



Numerous microorganisms require vitamins for growth, particularly 

 the members of the B group. Since different species and strains have 

 different requirements, a form can be selected that requires the factor 

 in question for normal growth. When all other necessary factors are 

 supplied in suitable amounts, the organism still does not multiply 

 imtil the vitamin to be assayed is added. Thereupon, the rate or the 

 total amount of growth depends upon the amount of the vitamin 

 added. Growth may be determined by microscopic counting, plate 

 counts, turbidity measurements, nitrogen determination on the washed 

 and collected cells, or in certain cases the titration of acids formed 

 in the medium by the growing culture. Methods are calibrated with 

 known amounts of pure vitamins. 



Many types of organisms are used in vitamin assays, and other 

 classes of compounds are determined in a similar way. Bacteria, fungi, 

 and yeasts are used to detect and measure vitamins, amino acids, 

 cations, and metabolic compounds of various kinds. 



To cite a typical example, the acid-producing bacterium Lactobacil- 

 lus casei will not grow normally if the nutrient medium is lacking 

 in folic acid. Control tubes containing media deficient in folic acid 

 are supplemented with graduated amounts of pure folic acid, inocu- 

 lated with a pure culture of L. casei, and incubated for a specified 

 period. The acid produced in each tube is measured by titration 

 with a standard alkali solution. Similar tubes containing graduated 

 amounts of the food to be tested are treated in a similar manner, and 

 the amount of acid produced is titrated. By comparing the acid pro- 

 duction in the experimental tubes with the acid production in the 

 control tubes containing pure folic acid, the amount of folic acid in the 

 food sample under examination can be estimated with speed and 

 accuracy. 



VITAMINS AND NUTRITION 



It appears that the point of diminishing returns is rapidly being 

 reached in the use of the classical approaches to the discovery of new 



