RELATIVE ACTIVITY OF AN EXZYME 



543 



temperature, (iii.) Filtered gastric juice or an artificial gastric juice made 

 from pepsin porci or from one of the above-mentioned commercial extracts. 

 Method. Take nine test tubes and put 1 c.c. of water into each except the 

 first. Into the first two tubes put J c.c. of the juice to be examined. Mix the 

 contents of the second tube by repeated sucking up into the pipette and blow- 

 ing out. Transfer 1 c.c. of the mixture to the third tube, from which, after 

 similar mixing, remove 1 c.c. and put it into the next tube, and so on, rejecting 

 the 1 c.c. removed from the ninth tube. The tubes now contain 1 c.c. of 

 fluid consisting of the juice of unknown strength diluted as follows : 



Place the tubes in a rack with a space between tubes 5 and 6. In this 

 space put a tube containing 1 c.c. of the prepared standard juice. To each 

 in turn add 1 c.c. of the calcium-chloride-milk mixture. This must be done 

 as rapidly as possible. Use a graduated 10 c.c. pipette for the purpose. The 

 operation should then take less than a minute to perform. Hold the rack of 

 tubes in the hand and gently tilt it occasionally, observing the way in which 

 the milk mixture flows on the sides of the tubes. Determine which tubes 

 show curd formation simultaneously with the standard tube. Suppose the 

 fourth tube curds a little before the " standard," which curds a little before 

 tube 5. The unknown juice, therefore, is between eight and sixteen times as 

 strong in rennin action as the standard. 



A new series of dilutions should now be made. Dilute the unknown juice 

 eight times, and with a graduated 2 c.c. pipette put the following amounts of 

 juice and water into six tubes. 



Tiihp No. 



Diluted juice (c.c.) 

 Water (c.c.) . 



0-4 

 1-6 



A control tube containing 2 c.c. of the standard enzyme solution is placed 

 near the middle of the series ; 2 c.c. of the milk mixture is added "as before 

 and the above procedure carried out. Suppose now the control tube and 

 tube 3 almost coincide in clotting time. If tube 3 is just a little earlier in 



Then 1-2 c.c. 

 = 2 c.c. of the 

 2/1-2 times as 



clotting than the control, take an interpolated value, 1-2 

 of a 1/8 solution of the unknown has an " enzyme strength " 

 standard strength. That is, the unknown juice contains 8 x 

 much enzyme as the standard = 13-3. 



Similar experiments may be carried out with other enzymes, using suitable 

 substrates. For example, determine the " strength " of the ptyalin in your 

 own saliva, using a diluted solution of Taka diastase as standard. 



In determining the strength of j)roteolytic and lipolytic enzymes a water 

 bath capable of containing the labelled tubes and of being maintained at a 

 con.stant temperature (38-40° C.) is essential. For substrate for the proteo- 

 lytic enzymes either the coagulated egg albumin suspension or the turbid 

 suspension produced by the precipitation of the serum proteins by sulpho- 



