514 11. LOCALIZATION OF THE SITE OF INHIBITION 



enzyme would tend to rise in concentration. Some consideration has al- 

 ready been given to the concentrations of intermediates in multienzyme 

 systems (Chai^ter 7) and it was i)ointed out that there are several factors 

 which may determine whether accumulation of an intermediate will occur 

 or not. If such an intermediate does indeed accumulate during inhibition 

 and this can be detected by analysis, it provides useful information on the 

 site of the block. Some examples of intermediate accumulation where this 

 X)henomenon has been of great importance in the study of the inhibitors 

 might be cited: the rise in hexose phosphates during inhibition by iodoace- 

 tate, the appearance of large amounts of keto acids following treatment with 

 trivalent arsenicals, the accumulation of citrate in fluoroacetate poisoning, 

 the rise in toxic aldehyde formed from alcohol in tissues subjected to di- 

 sulfiram (Antabuse), 'the increase in acetylcholine at synaptic junctions 

 when acetylcholinesterase is inhibited, and the progressive accumulations 

 of the catechol amines throughout the body after administration of the 

 monoamine oxidase inhibitors. These and other instances will be discussed 

 in greater detail in those chapters devoted to the inhibitors involved. 



Inhibition of an enzyme in a metabolic sequence is not invariably fol- 

 lowed by the accumulation of the substrate for this enzyme. There are 

 several iDossible reasons. An equilibrium may exist between the intermediate 

 whose metabolism is blocked and earlier intermediates in the sequence. 

 Thus triose phosphate does not rise appreciably during inhibition with 

 iodoacetate because the equilibrium conditions favor the hexose phosphates. 

 There may be other pathways for the metabolism of the intermediate 

 so that a block of one path will only shift the pattern of the process. Cer- 

 tain amines in certain tissues do not accumulate even though monoamine 

 oxidase is completely inhibited, presumably because reactions such as 0- 

 methylation are also available. Sometimes simple diffusion from the region 

 of formation will prevent the accumulation of an intermediate. The rel- 

 atively low permeability of mitochondria to citrate allows the marked 

 accumulation of this intermediate when aconitase is blocked, whereas other 

 cycle intermediates upon inhibition of their respective enzymes would 

 have a greater tendency to leave the mitochondria. It is thus often impor- 

 tant to analyze the medium or perfusate for the intermediate in such 

 cases and not confine the examination to the tissue units. It must also be 

 realized under certain circumstances that the accumulation of an inter- 

 mediate must be very great for it to be detected, especially when the steady- 

 state level is normally low. Thus a fair degree of inhibition may be exerted 

 without a demonstrable effect upon an intermediate. 



Conversely there are several reasons why the accumulation of an inter- 

 mediate during inhibition does not necessarily imply a block of the enzyme 

 metabolizing that intermediate. This is the case, as we have seen above, 

 when the intermediate is in equilibrium with other substances, and may be 



