New York Agricultural Experiment Station. 



283 



250 cc. of Jj- acetic acid, the liquid was siphoned off, the casein 

 washed several times with water, placed in a linen bag and squeezed 

 as dry as possible. It was then transferred to a mortar, ground to 

 a paste with a little water, the paste put into a flask and 150 cc. 

 of water, 75 cc. of lime water and some toluol were added to it. 

 After considerable shaking the lime water became saturated with 

 casein. By this process a solution was obtained containing a cal- 

 cium caseinate neutral to litmus but acid to phenolphthalein, 8 and 

 containing four equivalents of base. The undissolved casein was 

 removed by centrifuging and filtering. The amount of casein in 

 solution was determined and the solution so diluted that each 50 

 cc. contained 1 gram of casein. Fifty cc. portions of this solution 

 were withdrawn at intervals and precipitated with acetic acid. The 

 amounts of nitrogen found in the filtrates were as follows: • 



30 

 min- 

 utes 



5 

 hours 



12 



hours 



24 

 hours 



Milligrams of nitrogen in original solution 



Milligrams of nitrogen not precipitated by acetic 

 acid 



158 

 0.07 



158 

 0.92 



158 

 1.96 



158 

 2.00 



Casein solutions prepared in this manner gave the following 

 reactions. They were not coagulated by rennin. The addition of a 

 few drops of a 10 per ct. solution of calcium chloride caused them 

 to curdle; 9 the addition of one drop caused no change but the sub- 

 sequent addition of rennin produced coagulation. If enough hydro- 

 chloric acid was added to change the caseinate to one containing 

 two equivalents of calcium, 10 the addition of rennin caused coagu- 

 lation. That this coagulation was not due to the calcium chloride 

 formed by the acid was shown by the fact that rennin caused coagu- 

 lation after all this calcium chloride had been removed by dialysis. 

 In both instances the coagulation removed all the nitrogen from the 

 solution, as is shown by the following figures: 



Milligrams nitrogen in 

 original solution 



316 

 316 

 316 



Milligrams nitrogen not precip- 

 tated by rennin 



0.8 

 0.6 

 0.2+ 



The behavior of such caseinate solutions toward rennin can be 

 explained by the work of Van Slyke and Bosworth as follows : 



8 Robertson: Jour. Biol. Chem., 2: p. 317; Van Slyke and Bosworth: ibid., 14: 

 p. 211-225. 



9 Robertson: ibid., 2:381. Robertson believes that the addition of the common 

 Ca ion represses the dissociation of the caseinate and thus causes precipitation. 



10 Van Slyke and Bosworth: ibid., 14: pp. 211-225. 



