BASIC DIFFICULTIES IN TRACER METHODOLOGY 73 



whether or not exchange has occurred by allowing the precursors and 

 product to react under conditions where the biological or energy-produc- 

 ing system is interfered with. For example, animal tissues are known to 

 convert inorganic phosphate into phospholipid. This is probably not an 

 exchange process, because experimental observations showed that (a) 

 inorganic phosphate does not exchange with the phosphate radical of 

 phospholipid when sodium phosphate is shaken with a phospholipid solu- 

 tion, (b) homogenized liver failed to form radioactive phospholipid from 

 inorganic P^-, and (c) respiratory inhibitors such as cyanide and carbon 

 monoxide interfered with the formation of the tagged phospholipid (44). 



It will be recalled from Chap. 1 that the pickup of C* in the carboxyl 

 group of phosphoglyceric acid was interpreted to mean that this com- 

 pound was the first stable site of CO2 fixation in photosynthesis. It was 

 necessary, however, to show that this pickup is not merely due to an 

 exchange reaction. Fager and Rosenberg (45), by comparing simulta- 

 neously the kinetics of total fixation of CO2 and fixation in the phospho- 

 glyceric acid, were able to provide support for the postulation that the 

 labeling of PGA does indeed represent a step in the photosynthetic process 

 and not just an unrelated exchange reaction. 



A detailed listing of exchange reactions, particularly in organic systems, 

 has been compiled by Wahl and Bonner (11). It has been shown (22) 

 that no exchange occurs between inorganic phosphate ions in solution and 

 hexose monophosphate, glycerophosphate, lecithin, casein, nucleic acid, 

 pyrophosphate, and metaphosphate. No exchange was found between 

 the sulfur atoms of cysteine, thioglycolic acid, or urea and those of HoS, 

 S=, or SH~, respectively. In general, where the atom is tightly bound in 

 an organic molecule, there will be little opportunity for exchange (e.g., 

 hydrogen in benzene, carbon in the chain). 



Care must be taken to prevent the loss of the label from the molecule 

 under study. The 11+ of — OH, — COOH, — NHo, for example, is known 

 to exchange readily with the H+ of water. A tritium or deuterium label 

 in such a position would become widely distributed in an aqueous medium 

 and would no longer be indicative of the labeled molecule. A tritium or 

 deuterium label for an aldehyde might be stable under some conditions 

 but would be lost if the aldehyde underwent oxidation. 



Important considerations resulting from the exchange reaction that 

 occurs between certain ions in blood and those in the bone crystal have 

 been largely overlooked in the interpretation of studies of labeled calcium 

 and phosphate in animals. It is well recognized that fresh bone sus- 

 pended in a solution containing Ca^^ ions can incorporate these ions by the 

 process of exchange (46 to 48). Phosphate, strontium, and radium ions 

 can likewise be incorporated. Experiments have shown that 10 to 20 

 per cent of the calcium in bone is exchangeable, depending on the part of 



