422 PRACTICAL ORGANIC AND BIO-CHEMISTRY 



The fibrin is prepared in the following way : 



Fresh fibrin is minced, washed till free from blood and placed 

 for 24 hours in a 0*5 per cent, solution of congo red in the propor- 

 tions of 50 gm. of fibrin to 100 c.c. of congo-red solution. The mass 

 is poured into a large volume of water heated to 80 to fix the dye 

 and kept at this temperature for about 5 minutes. The fibrin is then 

 placed in a cloth and washed in running water ; the excess of water is 

 squeezed out and the fibrin is preserved in a mixture of equal parts of 

 glycerol and water, a little toluene being added as a preservative. 



Another modification consists in the use of stained cubes of 

 coagulated egg-white. 



A known quantity of 0*5 gm. of congo-red fibrin, 5 c.c. of 0*4 per 

 cent, hydrochloric acid and 5 c.c. of the pepsin solution are placed in 

 a test tube and put in a water-bath at 40 for half an hour. After 

 this time sufficient solid anhydrous sodium carbonate is added to 

 change the blue colour of the congo red to red. This also stops the 

 action of the enzyme. A measured volume is removed ; in order 

 to compare two pepsin digests water is added from a burette to the 

 deeper one till the tints of the two solutions are the same. The 

 amount of water added is noted. The strengths of the enzymes are 

 to one another as the amount of dilution: thus if an equal volume of 

 water be added the strengths are as 2 : I. 



(d) FulcCs Method. 



By making use of edestin as substrate and its precipitability by 

 salts from its solution in hydrochloric acid, Fuld has devised a very 

 simple method for measuring the activity of pepsin solutions. Vary- 

 ing amounts of the enzyme are added to definite volumes of the 

 edestin solution in a series of tubes and after a prescribed lapse of 

 time sodium chloride is added ; the first tube in each series in which 

 a precipitate of edestin is no longer formed is noted, i.e. the tube con- 

 taining least enzyme. Thus : 



5 c.c. of the 0-5 per cent edestin solution in 0*4 per cent, hydro- 

 chloric acid are measured out with a pipette into each of a series of 

 five test tubes. To these tubes is added in order, 0-2 c.c, 0*4 c.c., 

 0*6 c.c., O'8 c.c., i-o c.c. of the pepsin solution A from a burette 

 (generally 2 drops = O'l c.c.). The same operations are performed 

 with pepsin solution B. 



The tubes are kept at the ordinary temperature for half an hour, or 

 longer, but the same time for each series ; then to each tube is added 

 I c.c. of saturated sodium chloride solution. The first tube in which 

 a precipitate of edestin is no longer produced is noted. 



