ANIMAL FLUIDS AND TISSUES. 439 



Leo's method. This is employed for very accurate determinations 

 of hydrochloric acid. It is based upon the principle that free acids 

 are fully neutralized by the addition of calcium carbonate even at 

 the ordinary temperature, while solutions of acid phosphates or of 

 other acid salts retain their acidity. 



To 10 c.c. of gastric nitrate are added 5 c.c. of concentrated solu- 

 tion of calcium chloride and a few drops of phenol-phtalein ; titra- 

 tion is made with deci-normal potassium hydroxide (result A). To 

 15 c.c. of a second portion of gastric nitrate is added 1 gramme of 

 pure, powdered calcium carbonate ; the mixture is well shaken and 

 filtered through a dry filter. Ten c.c. of this filtrate are removed, and 

 air is passed through it in order to remove all carbon dioxide, which 

 interferes with the use of phenol-phtalein as an indicator. (A 

 double-bulbed syringe, to one end of which a piece of glass tubing is 

 attached, answers well for this purpose.) Having added to the 

 filtrate freed from carbon dioxide 5 c.c. of calcium chloride solution 

 and phenol-phtalein, the titration is made (result B). As titration 

 A gives the total acidity, titration B the acidity due to acid salts, 

 therefore, A-B equal the alkali used for neutralizing the free acids. 

 If fatty and lactic acids are not present the result indicates hydro- 

 chloric acid. Should these acids be present, they must first be 

 removed the fatty acids by distillation, the lactic acid by agitation 

 with ether. 



g. Pepsin and pepsinogen. In case free acid is present, 10 c.c. of 

 gastric juice are placed in a beaker, and a small bit of dried fibrin, 

 or a lamella of blood albumin (Merck), is added, and the beaker 

 placed in a thermostat at a constant temperature of 38 to 40 C. 

 (100 to 104 F.). Pepsin is indicated by the rapid solution of the 

 flake of albumin. If free hydrochloric is absent, the juice is rendered 

 acid with a drop of this acid and then tested in the manner described. 



h. Rennet ferment and rennet zymogen. Rennet is tested for as fol- 

 lows : Ten c.c. of gastric juice are exactly neutralized with deci-normal 

 alkali and mixed with an equal volume of neutral unboiled, or better 

 boiled, milk. The mixture is placed in a thermostat at 38 C. (100 

 F.). If a casein coagulum is formed in ten to fifteen minutes, the 

 coagulation is due to the rennet ferment. 



Rennet zymogen is detected thus : Ten c.c. of gastric juice are 

 rendered feebly alkaline and mixed with 2 c.c. of a 1 per cent, solu- 

 tion of calcium chloride and 10 c.c. of milk. If the rennet zymogen 

 be present, a heavy cake of casein is precipitated in a few minutes. 



*. Detection of proteids. Of these, syntonin, albumoses, and pep- 



