44 RADIOISOTOPES IN BIOLOGY AND AGRICULTURE 



to be directly related to the two-carbon acceptor and is probably the 

 immediate precursor for it in a cleavage into a three-carbon compound 

 and a two-carbon compound. 



Biosynthesis. Labeled organic compounds may be prepared by chem- 

 ical synthesis or biosynthesis. Since many of the important biological 

 metabolites are exceedingly complex, the biological synthesis is often 

 most convenient. The general procedure is to grow the biological mate- 

 rial under such conditions that the radioactive label enters into the metab- 

 olism and becomes incorporated into the various metabolites produced. 

 One consideration is that with biological synthesis the label usually 

 becomes randomly located at all possible sites. This is especially true 

 for radioisotopes of carbon or hydrogen. However, random labeling is 

 not usually a problem with elements such as phosphorus, cobalt, sulfur, 

 or iodine which are most often located in specific positions in the molecule. 



The bibhography in reference (100) may be consulted for a listing of the 

 various labeled compounds that have already been synthesized biolog- 

 ically. Plants, animals, and microorganisms have been utilized for this 

 purpose. Some of the more important types of compounds labeled 

 include sugars from tobacco and crop plants; drugs from medicinal plants; 

 amino acids, nucleic acids, and vitamins from bacteria, yeasts, and molds; 

 fatty acids and pigments from algae; and insulin, cortical steroids, choles- 

 terol, and glycogen from tissue cultures. 



These procedures may also be employed to determine whether or not a 

 given element is a constituent of a particular metabolite. For example. 

 Totter et al. (101) showed that molybdenum was a nondialyzable compo- 

 nent of xanthine oxidase by administration of radiomolybdenum to a ' 

 dairy cow and subsequent isolation of the enzyme from the milk. In all 

 the purification steps the ratio of radiomolybdenum to xanthine oxidase 

 did not change; also, when the isotope was merely added to milk, it did 

 not enter the enzyme. From these experiments it was also possible to 

 calculate that there was a molar ratio of flavin to molybdenum of 2:1. 

 The biosynthesis of a vitamin-Bi2-like compound in both the gastro- 

 intestinal tract and the tissues of sheep was demonstrated by Monroe 

 et al. (102). This was done by administration of inorganic labeled cobalt 

 to the animals and subsequent fractionation of the tissues and excreta 

 for radioassay. 



ADVANTAGES OF DOUBLE LABELING 



The simultaneous use of two radioisotopes in an experiment is made 

 feasible by the fact that measurements of each in the same sample are 

 possible (see Chap. 5). In general, double labeling offers a means of 

 comparing directly in a biological system the movement, synthesis, or 



