206 THE HYDROLYZING ENZYMES 



Thus several of the oxidizing enzymes, or Oxidases regain their activity, 

 lost by heating, when the solution is allowed to stand for some time at 

 ordinary temperatures. According to Gramenetski the same phe- 

 nomenon may be displayed by certain Diastases or starch-splitting 

 enzymes and even in some measure by Trypsin. The vegetable 

 Proteases or protein-splitting enzymes sometimes withstand higher 

 temperatures than the corresponding enzymes of animal origin and 

 Karl Meyer has drawn attention to the rather extraordinary fact that 

 the Trypsin which is produced in culture-media by Bacillus prodigiosus 

 will withstand heating for fifteen minutes to 100 C, although it is 

 destroyed within thirty minutes at 56 C. This looks rather as though 

 a trypsin-splitting ferment also existed in the culture-medium for 

 which the optimum temperature is about 56, and which is destroyed 

 by higher temperatures more rapidly than trypsin itself. 



It would, therefore, be very unsafe to infer because a substance does 

 not lose its characteristic activity of some type or other when it is 

 heated that it is therefore not an enzyme. It would be still more 

 unsafe, of course, to infer that it is an enzyme simply because it is 

 "inactivated" by heat. Yet both of these inferences, unfortunately, 

 have not infrequently been made in biological and biochemical investi- 

 gations. In deciding whether or not a substance or material should 

 be classed as an enzyme we should be guided, rather, by its quantitative 

 relationship toward the particular activity which it displays. The 

 enzymes are usually effective in relatively minute concentration. It 

 has been estimated, for example, that a certain Rennin or milk-coagulat- 

 ing enzyme preparation will convert no less than 500,000 times its 

 weight of casein into the coagulating form, paracasein, and a prepara- 

 tion of pepsin has been obtained which will hydrolyze to peptones 

 100,000 times its weight of fibrin. The excessive amount of change 

 which may thus be brought about by relatively minute proportions of 

 enzymes almost compels the assumption that they are not consumed 

 during the progress of the changes which they accelerate, for otherwise 

 the enzyme would probably be "used up" long before so immense a 

 proportion of change had been accomplished. It is, however, impos- 

 sible at the present stage of our knowledge to submit this supposition 

 to vigorous investigation, because the various hydrolyzing enzymes, 

 at all events, are themselves chemically unstable substances and 

 undergo spontaneous transformation resulting in inactivation on 

 standing in aqueous solution. They are carried down together with 

 any precipitates which may be formed in the digestion-mixture and 

 are not infrequently partially bound by or combined with not only the 

 substrate, but also the products of hydrolysis. Any chemical procedure 

 designed to isolate and recover the enzyme from a digest in which it has 

 been operating would involve loss or impairment of the enzyme even 

 if it had been dissolved in distilled water instead of in a solution of the 

 substrate which it attacks and of the products of its hydrolysis. No 



