PROTEOLYTIC ENZYMES UPON GELATIN 187 



EXERCISE 12. TO SHOW THE ACTION OF PRO- 

 TEOLYTIC ENZYMES UPON GELATIN 



Apparatus. Phenol, 0.5% solution (10 c.c. of 5.0% 

 phenol +90 c.c. distilled H2O) ; water bath and thermometer; 

 gelatin, 7 gms.; 15 tubes sterile gelatin; formalin; xylol; 

 5 sterile 1 c.c. pipettes; centimeter scale. 



Cultures. B. ramosus; B. fluorescens; B. subtilis; 

 B. mycoides; B. prodigiosus. 



Method. 1. Make two gelatin stab cultures of each 

 organism and when nearly all liquefied (2-5 days old) pro- 

 ceed with the experiment. 



2. Dilute the 5% phenol to 0.5% as above, with distilled 

 water. 



3. Add 7 gms. gelatin, dissolving by heating not over 

 70 C. Neutralize carefully. 



What is the source of acid in phenol gelatin? Why is 

 the gelatin neutralized? 



4. Select five test tubes having the same diameter. Fill 

 each half full. Solidify in an upright position. 



5. Shake each of the liquefied cultures with 3 to 4 c.c. 

 of xylol. 



6. After one hour, add 1 c.c. of the clear supernatant 

 xylol solution of each culture to a tube each of solid phenol 

 gelatin and of ordinary gelatin. With a blue pencil, mark 

 the surface of the solid gelatin. 



7. Examine the tubes daily. Is there any evidence of 

 growth? Of liquefaction? If liquefaction is noted, measure 

 its progress in millimeters. 



8. Save the original cultures with which the xylol 

 has been shaken. Is there any evidence of further growth? 



9. If 1 c.c. of a liquefied gelatin culture of a liquefying 

 organism were added to a tube of solid ordinary gelatin, 

 what would happen? What would result if it were added 

 to a tube of solid phenol gelatin? Explain. 



10. What action does the xylol have? How can you 



