230 INACTJVATION OF TRYPSIN. I 



Experimental Procedure. 



Preparation of the Gelatin. — Salt-free gelatin was prepared as described by 

 Loeb,® i.e., the gelatin was brought to the isoelectric point and then washed 

 (except that 100 gm. were prepared at a time). The gelatin was melted, diluted 

 to about 5 per cent and titrated to a pH of 6.3 with NaOH. It was then diluted 

 so as to contain 2.5 gm. dry weight per 100 cc. and sufificient KCl added so that 

 the resulting solution had a specific conductivity at 33°C. of 2 X 10~^ reciprocal 

 ohms. A few crystals of thymol were added and the solution was kept in the 

 ice bo.x. 



Preparation of the Trypsin. — The trypsin used in all the experiments was a 

 sample of Fairchild's trypsin. It was prepared for use by suspending 5 gm. in 

 50 cc. of water and dialyzing under pressure at 6°C. for 18 hours. The solution 

 was then filtered from the rather heavy precipitate and sufficient KCl added to 

 bring the specific conductivity to 2 X 10 ~^. This solution is very unstable and 

 loses its activity quite rapidly even at 3°C. It was prepared fresh each day. 



Determination of the Conductivity. — The apparatus used was a Leeds and 

 Northrup Kohlrausch bridge and resistance box. The change in the bridge 

 readings were used direct to avoid calculation. These readings are related to 

 the actual conductivity by the formula 



^^^1000-. 4^ 



in which .Y = resistance of solution, A = bridge reading, and R = resistance 

 of standard resistance box. It will be seen that if the readings are always 

 begun at the middle of the bridge (500), the first ten or fifteen points will 

 each represent very nearly equal changes in the conductivity and may be 

 considered as proportional to the percentage change in the conductivity. 

 Since the conductivity increases the bridge readings will decrease. If all the 

 solutions have the same conductivity at the beginning of the experiment and the 

 resistance is so chosen that the bridge reading at the beginning is 500 in each case, 

 then a change in the bridge reading of from 500 to 490 will represent the same 

 change in the conductivity of each of the different solutions. 



Type of Conductivity Cell. — The cell used is shown in Fig. 1. Fifteen such 

 cells were made and adjusted (by warming the cement and moving the electrode) 

 so that they all had the same constant, 3.5. The electrodes were plated with 

 platinum black. Readings could be made with ease to half a scale division on 

 the bridge. As little current as possible was run through the cell, although no 

 effect due to the passage of current could be noted. 



pH Determinations. — The determinations were made by the e. m. f. method. 



Formol Titration. — The titration was carried out as described in a previous 

 paper!' by adjusting the solution to pH 7.0 with neutral red as an indicator 



»Loeb. J., J. Gen. Physiol., 1918-19, i, 237. 

 ^0 Northrop, J. H., /. Gen. Physiol, 1919-20, ii, 595. 



