462 ENZYMES 



In addition to purification by selective adsorption, selective 

 elution sometimes may be effective. Thus y-alumina will 

 adsorb both saccharase and maltase, both of which are liberated 

 by weakly acid or neutral phosphate solution. A primary 

 phosphate, however, will liberate saccharase completely, most 

 of the maltase remaining behind, and may be recovered by 

 elution with a secondary phosphate. 



The adsorption method of purification tends to show that 

 what was hitherto supposed by some to be responsible for the 

 catalytic action are removable impurities. Thus Willstatter 

 and his collaborators found that they were able to reduce 

 the iron content of their peroxidase preparations from 0-5 to 

 006 per cent without loss in activity. Similarly it has been 

 shown that phosphorus is not responsible for the activity nor 

 is an essential constituent of saccharase which, by adsorptive 

 methods, has been freed almost completely from carbohydrate, 

 protein, and phosphorus without loss of stability or activity. 



Owing to the absence of chemical criteria for determining 

 the effect upon enzymes of the process of purification, special 

 methods have been devised for ascertaining the alteration 

 in activity of an enzyme in the course of its purification ; 

 some of these methods are outlined under the respective 

 enzymes (see under Lipase and Peroxidase). The criterion 

 suggested by Euler and Josephson for saccharase * which has 

 been adopted by Willstatter, is known as the " Time Value " 

 (Zeitwert) ; this is the time in minutes required by 0-05 gram 

 of the enzyme preparation dissolved in 5 c.c. of i per cent 

 sodium phosphate and added to 20 c.c. of 20 per cent sucrose 

 to reduce the rotation of the solution to zero, the temperature 

 being 15-5° C. By a number of alternate adsorptions and 

 elutions, including the use of lead phosphate, Willstatter f 

 has prepared from yeast a saccharase preparation having a 

 " time value " of o-i, compared with a value of 300 for the 

 yeast from which he started, which implies a 3000-fold in- 

 crease in activity. The purest product is free from carbo- 



* Euler and Josephson : " Zeit. physiol. Chem.," 1925. I45» 130- 

 t Willstatter : id., 1926, 151, i; " Annalen," 1922, 427, iii, and 

 earlier papers. 



