46 THE GENERAL CHARACTERS OF THE PROTEINS 



The method has been applied by Hardy (see below, p. 53) for 

 investigating the amount of hydrolysis which takes place in dilute 

 solutions after a protein has been neutralised, the neutralisation 

 point having been determined by other methods. 



The method is of somewhat limited application, owing to the fact 

 that most proteins are hydrolysed to some extent even by dilute acids. 



B. I. Direct Titration in the Presence of Indicators. 



This method is quite effective in the case of strong bases, such 

 as the protamines. It requires to be applied with some care in the 

 majority of cases, owing to a variety of circumstances, such as the 

 hydrolysis of the salts in dilute solutions, and the capacity for the 

 formation of acid and basic salts. Particular stress must also be 

 laid upon the choice of indicators. The method is discussed in 

 some detail below, in considering the researches on certain individual 

 proteins. 



B. II. The Determination of the Solubility of Proteins which are 

 Insoluble in Water, in Acids and Bases. 



This method is applicable to a limited number of proteins only, 

 e.g., to the globulins. This is also discussed in greater detail below. 



B. III. The Determination of the Acidity of the Filtrates from 

 Protein Precipitates Produced by Neutral Salts or Alkaloidal 

 Reagents. 



This method has been employed by Spiro and Pemsel, Cohn- 

 heim and Krieger, and von Rhorer. 



Spiro and Pemsel added acids and alkalis in excess to protein 

 solutions. They then precipitated the proteins by ammonium sul- 

 phate, and estimated the acid or base remaining in the filtrate. They 

 assumed that the combination of acid or base with protein could be 

 precipitated by salts in the same way as the protein itself, and arrived 

 thus at conclusions as to the amount of acid or base which could 

 enter into combination with proteins. This method has been but 

 little employed, owing to the difficulty of maintaining solutions of 

 ammonium sulphate in a state of complete neutrality. 



Cohnheim and Krieger employed a similar method, using, how- 

 ever, alkaloidal reagents, assuming that the acid salt of the protein 

 and the reagent enter into double decomposition, according to such 

 an equation as the following : 



Protein hydrochloride + Calcium phosphotungstate = Protein phosphotungstate + CaC! 2 . 



If to protein containing excess of acid, calcium phosphotungstate 

 be added, the total acidity of the solution, as determined by titration, 

 would be diminished, owing to the combination of acid and protein, 

 and subsequent double decomposition of the salt thus formed with 

 the alkaloidal reagent. Other reagents used were sodium picrate, 

 calcium trichloracetate and potassium mercuric iodide. 



The method of Cohnheim and Krieger was subjected to a critical 

 examination by von Rhorer, who has shown that, as originally 

 carried out, it is not free from certain errors. 



When, for example, calcium phosphotungstate is used as precipi- 



