SELMAN A. WAKSMAN 271 



used lately with great success. The solution of the enzyme is filtered through paper 

 or through a Charnberland or other convenient filter. Salts can also be removed by 

 dialysis. The enzyme can be precipitated from solution by alcohol or by a definite 

 concentration of a sulphate, preferably ammonium sulphate. The use of different 

 concentrations of salt frequently permits the separation of one enzyme from another. 

 The precipitation is followed by selective adsorption and elution of the enzyme. Posi- 

 tively charged aluminum hydroxide and negatively charged kaolin have been used 

 extensively as adsorbing agents. 



The microbial cell begins to liberate the enzyme into solution only after its death, 

 as in the case of yeast. The autolysis of the cells or the enzymatic degradation of the 

 protoplasm of the microbial cell is accompanied by the passing of the various enzymes 

 into solution. Gentle autolysis accompanied by adsorption processes resulted in the 

 liberation of much more active enzymes than previously obtained.' 



For the concentration of bacterial enzymes, the following method has been rec- 

 ommended:^ Pure cultures of bacteria are grown in i liter portions of nutrient media 

 for three days. The culture is then treated with an aqueous emulsion of mastix and 

 acidified with acetic acid. The emulsion is prepared by dissolving mastix in alcohol 

 and diluting with an equal volume of water. The precipitate which is formed as a 

 result of treatment of the culture is allowed to stand twenty-four hours and is then 

 filtered off. The mastix is removed from the precipitate by dissolving with alcohol 

 and ether. The residual precipitate contains the bacterial enzymes. 



MEASUREMENT OF ENZYME ACTION 



An enzyme is characterized not by its structure, which is so far unknown, but 

 by its activities. The methods of demonstrating enzyme action are based either upon 

 the disappearance of the specific substrate or the formation of reaction products. The 

 action of diastase (amylase) upon starch is measured, on the one hand, by the dis- 

 appearance of starch, as shown by the iodine reaction, or by a change in the colloidal 

 condition of the starch, and, on the other hand, by the formation of dextrins and 

 maltose, etc. Unfortunately, we are dealing in most instances with mixtures of en- 

 zymes and not with pure enzymes. Hence it is frequently difficult to determine just 

 which products are formed by one enzyme and which are acted upon by another. 



INFLUENCE OF ENVIRONMENTAL CONDITIONS 



Enzymes are influenced in various ways by physical and chemical factors, which 

 either favor (activate) or injure (paralyze) their action. 



Increasing temperatures, up to a certain region, bring about an increase in en- 

 zyme action, according to the laws of chemical kinetics; however, even at relatively 

 low temperature an injurious action sets in. Most enzymes are very sensitive to 

 temperatures above 70° C, even during a very brief period. Temperatures above 

 45° C. prove injurious after a prolonged period of action. The temperature coefficients 

 (Arrhenius' constant A) become smaller with increasing temperatures, finally be- 

 coming negative; different enzymes, however, behave differently. The optimum tem- 



' Willstatter, R.: op. cit., p. 1359. 1927. 



' Schierge, N.: Biochem. Ztschr., 179, 248. 1926. 



