CASEIN 91 



error lying in the fact that after the attainment of neutrahty to 

 litmus additional casein dissolves in the solution with extreme 

 slowness. Many hours of rapid shaking or stirring are quite 

 insufficient, at room temperature, to bring about complete "satu- 

 ration" of the alkali with casein. Resource was had, therefore, to 

 the device of dissolving weighed amounts of casein in measured 

 volumes of alkali of known concentration, and then neutralizing 

 the excess of alkali by the addition of standardized acid, stirring 

 vigorously the while, until the casein just began to be precipitated. 

 The point at which precipitation began was readily determined, 

 with considerable accuracy, by the change (diminution) of the re- 

 fractive index of the solution. In this way it was ascertained that 

 at complete "saturation" of an alkali (NaOH or LiOH) one gram 

 of casein binds 11.4 X 10~^ equivalent gram molecules of the base. 



These determinations have been repeated by Van Slyke and 

 Bosworth (57), whose results very closely coincide with mine. 

 They find that at the point at which further diminution of the 

 proportion of base to casein results in the precipitation of free 

 casein one gram of casein is combined with between 11.0 X 10~^ 

 and 11.5 X 10~^ equivalents of ammonium, sodium or potassium. 



These "saturated" compounds of casein with bases react in 



solution to indicators as follows (41): 



Dimethylaminoazobenzol yellow 



Congo red red 



Sodium alizarinsulphonate red 



Cochineal rose 



Paranitrophenol colorless 



Rosolic acid yellow 



indicating, according to the determinations of Salm (48) an acidity 

 (H+ concentration) of about 10~^. According to Michaelis and 

 Rona (26) and Allemann (1), the optimum reaction for the complete 

 precipitation of casein is 2 X 10"* H + . 



It is not possible with equal simplicity of technique to determine 

 the combining capacity of casein for the alkaline earths at "satura- 

 tion " of these bases with casein, because the chloride of the alkaline 

 earth, which is necessarily formed in the solution at the same time, 

 precipitates the caseinate.* Thus four grams of casein were dis- 



* The "neutral" and "basic" caseinates of the alkaline earths are likewise 

 precipitated by the addition to their solutions of a somewhat greater propor- 

 tion of the corresponding alkaline earth. Cf. A. S. Loevenhart (24) and Van 

 Slyke and Hart (59). 



