182 Report OF THE CHEMIST OF THE 
added 2.5 grams of sodium chloride, thus making a 5 per ct. 
salt-solution. On warming, the unsaturated casein chloride 
completely dissolved. The experiment was repeated several 
times with a uniform result. On adding more acid to the salt- 
solution, the proteid reprecipitated as the saturated chloride 
of casein. Casein behaved toward acetic acid in the same 
manner, 
(2) Determination of amount of 7, hydrochloric acid required 
to saturate casein.— Fresh milk was drawn from a cow directly 
into a bottle containing ether, thus checking all acid fermenta- 
tion. We diluted 5 cc. of this milk, containing .0261 grams of 
nitrogen, with 50 ce. of water that had been boiled free from 
all carbon dioxide. We warmed the mixture to 104° F. (40° C.) 
and added 10 ce. of ;% hydrochloric acid. It was desired now 
to determine the amount of hydrochloric acid that had not 
combined with casein. Determinations of the uncombined acid 
did not give uniform results, when the titration was made 
directly after the addition of acid. We therefore employed the 
method of using calcium picrate." We added 5 cc. of a neutral 
solution of calcium picrate to precipitate all proteids com. 
pletely. The precipitate was filtered, and the filtrate titrated 
with 7, sodium hydroxide, using rosolic acid as an indicator. 
The reactions taking place can be represented, theoretically, 
as follows: 
Casein (O I),+ 3HCI (excess)=Casein Cl, + 2H,O + HCI. 
Casein Cl,+Ca picrate + HCl=Casein picrate + CaCl,+ 11Cl. 
When calcium picrate is used, the chlorine which was com- 
bined with casein is changed into calcium chloride and, in this 
form, does not interfere with titration by alkali. Thus, we 
are able to determine more satisfactorily the amount of acid 
left uncombined with proteid. 
In the experiment employed, it required 7.4 cc. of ,5. sodium 
hydroxide to neutralize the excess of acid, indicating that 2.6 
ce. of acid had entered into combination with .0261 grams of 
proteid nitrogen. Using 6.39 as the nitrogen factor of casein, 
0261 gram of nitrogen is equivalent to 0.167 gram of casein, 
and this amount combined with 2.6 ce. of ; hydrochloric 

“Rohrer. Archiv f. Physiol., 90: 368 (1902). 
