308 PRACTICAL PHYSIOLOGY 



This method, when congo red is used, serves for the rough estimation of 

 both gastric and pancreatic juices. Place an equal amount (weighed) 

 of stained fibrin 1 into two test tubes, and according to the ferment 

 under investigation add 15 c.c. 0-4 per cent. HC1 or 0-4 per cent. 

 Na 2 CO 3 solution to one, and to the other 10 c.c. 0-4 per cent. HC1 or 

 0-4 per cent. Na 2 CO 3 solution and 5 c.c. of the gastric or pancreatic 

 juice to be tested. Mix well and place in a water bath or incubator 

 at 40 C. for fifteen to twenty minutes. Filter off undigested fibrin 

 and examine the filtrate. Or if preferred two samples of digestive 

 juice of different strengths may be compared. The more active the 

 juice the more pigment in solution. The colour in the case of gastric 

 digestion is bluish and is difficult to compare ; the red colour can be 

 restored if the solution is rendered just alkaline by the addition of a 

 very small amount of solid sodium carbonate. 



Other methods have been introduced for the quantitative estima- 

 tion of both pepsin and trypsin in which the amount of digestion 

 of proteins like edestin, ricin, egg albumen and caseinogen is deter- 

 mined. The test with caseinogen is perhaps most readily carried out 

 as the material required is most easily obtained ; 1 gm. of com- 

 mercial casein is dissolved in 16 c.c. of 25 per cent. HC1 (sp. gr. 

 1-124) in a litre flask on the waterbath and then made up to 

 1,000 c.c. with water ; 10 c.c. of this solution are placed in each of a 

 series of test tubes and then varying amounts from 0-1 to 1 c.c. of the 

 gastric juice to be tested are added. The tubes are then placed in a 

 waterbath at 40 C. and allowed to digest for fifteen minutes. To 

 each test tube there is then added a few drops of a 20 per cent, solution 

 of sodium acetate ; this precipitates all undigested caseinogen. The 

 first tube in which a mere trace of clouding is observed is taken as 

 containing the amount of enzyme which just sufficed to complete the 

 digestion. The result is usually stated in terms of units. The reciprocal 

 of the amount which just suffices to complete digestion gives a measure 

 of the proteolytic activity of the juice, e.g. in a test if 0-03 c.c. of juice 



sufficed to complete the digestion the value would be put as -^ or 33 



units. 



When trypsin is to be determined the caseinogen (0-1 gm. ) is dissolved 



N 

 in 5 c.c. -JQ NaOH, 25 c.c. water and then solution is boiled. After 



N 

 cooling the alkaline solution is neutralised with y^ HC1 and the volume 



made up to 100 c.c. (This solution does not keep well and ought 

 always to be freshly prepared.) A series of test tubes, as before, are 

 filled with 2 c.c. caseinogen solution, varying volumes of trypsin 

 solution or pancreatic juice added. At the end of the period of digestion 

 (one hour) caseinogen not digested is precipitated by means of acid 

 alcohol (1 c.c. glacial acetic acid, 49 c.c. water and 50 c.c. 95 per cent, 

 alcohol). End point and calculation as above. 



Mett's Method. A narrow glass tube (1-2 mm. diam.) drawn to a 



1 Carefully cleaned minced fibrin is allowed to soak in 0-5 per cent, congo 

 red solution for twenty -four hours (50 gms. moist fibrin per 100 c.c. solution). 

 It is then poured into a large volume of water and heated for five minutes to 

 80 C. It is then collected on a cloth, well washed under the tap, squeezed 

 as dry as possible and kept in equal parts glycerine and water. Add a little 

 toluol as preservative. 



