THE REPORTING OF INHIBITION STUDIES 885 



(5) Do inhibitor incubation experiments when indicated. It is very useful, 

 especially with a new inhibitor, to incubate the enzyme with the inhibitor 

 for varying times and under various conditions. Not only will this provide 

 important information on the rate of inhibition but can well point to the 

 mechanism of the inhibition. The appropriate techniques are discussed 

 in Chapter 12. 



(6) Determine the degree of reversibility of the inhibition. The reversibility 

 of an inhibition will not only give information of a practical nature that 

 can be useful in future work with the inhibitor, but also can indicate in- 

 hibition mechanisms, particularly if a chemical reaction with the enzyme 

 is involved. It also serves to demonstrate whether secondary inactivation 

 of the enzyme or damage to the cells has occurred during the inhibition. 



(7) Exercise caution in the interpretation of protection or reversal experi- 

 ments involving substances reacting ivith the inhibitor. The protection of 

 an enzyme or a cell preparation from an inhibitor by a substance reacting 

 with or complexing with the inhibitor is not very significant and does not 

 indicate the mechanism by which the inhibition occurs. In such cases, one 

 is merely reducing the concentration of free or effective inhibitor. Rever- 

 sal experiments may be more informative, but only if analyzed quantita- 

 tively and correctly. At the most, such experiments only show the relative 

 affinities of the inhibitor for the enzyme and the protector or reversor. 



SUGGESTIONS RELATIVE TO THE REPORTING OF 

 INHIBITION STUDIES 



(1) Report the experimental conditions conrpletely. It is necessary to state 

 accurately "all of the conditions under which the inhibition experiments 

 were run. Some factors that may seem unimportant when the report is 

 written may later be of major significance. Particularly in interpreting 

 early work, it is often very difficult to determine in what manner the ex- 

 periments were conducted. In studies with purified enzymes the following 

 would be the minimally required information: {a) pH, (b) temperature, 

 (c) ionic composition of the medium, (d) the gas phase in equilibrium with 

 the medium, (e) the substrates and their concentrations, (/) other substances 

 present, such as buffers, stabilizers, cofactors, etc., (g) the exact source of 

 the enzyme, (h) the intervals of time during which the inhibition was de- 

 termined, and (i) how the rates were determined. In certain instances, 

 other conditions, such as ionic strength, may be important. Studies on cells 

 or tissues require in addition a complete history of the material, including 

 nutrition of the cells before the experiment, their age, their growth rate 

 or state of functional activity, and any other pertinent information. Un- 

 explained discrepancies between the data of different investigators often 

 result from deficiencies in reporting. 



