152 COMPOUNDS OF THE PROTEINS 



Amphoteric character of the proteins. A variety of facts have been 

 ascertained in recent years, however, which have compelled a revision 

 of this opinion, and we now recognize that some elements in the protein 

 molecule other than free NH 2 or COOH groups must be responsible 

 for the acid- and base-neutralizing power that is possessed in a marked 

 degree by many proteins. 



In the first place, the investigations to which reference has been 

 made above have shown that only a very small proportion of the 

 nitrogen in proteins is present within their molecules in the form of 

 NH 2 groups. Thus, in the case of Edestin, as the above-quoted 

 estimations show, only 1.8 per cent, of the total nitrogen is present in 

 the form of NH 2 groups. Now edestin is insoluble, when in the free 

 condition, in water. It forms an insoluble hydrochloride containing 

 14X10" 5 equivalents 1 of hydrochloric acid per gram. On further 

 addition of acid, soluble hydrochlorides are formed, and the substance 

 passes completely into solution when the proportion of combined acid 

 is just double that contained in the insoluble hydrochloride. Still 

 further additions of acid, however, continue to be neutralized by the 

 protein until, at neutrality to tropaeolin OO, which changes color when 

 the amount of free acid in solution is between one-hundredth and one- 

 thousandth normal, after due allowance for the acid which remains 

 unneutralized, it is found that edestin combines with no less than 

 127 X 10~ 5 equivalents of acid per gram. The formation of the insoluble 

 compound with 14XlO~ 5 equivalents of acid must correspond to the 

 union of at least one molecule of the acid with each molecule of protein, 

 for any acid in excess of this amount results in the formation of a 

 compound of quite a different character, namely, one which is soluble 

 in water. The maximal number of molecules of acid which may be 

 neutralized by a molecule of edestin has not been determined, but it 

 evidently cannot be less than the number corresponding to 127X10" 5 

 equivalents of hydrochloric acid per gram of protein. If we assume, 

 therefore, that the insoluble compound represented the result of union 

 of one molecule of acid with each molecule of protein, then the com- 

 pound formed at neutrality to tropaeolin must represent the formation 



of a compound containing -T-T- = 9 molecules of acid for each molecule of 



edestin. If this compound were formed by the union of the acid with 

 NH 2 groups in accordance with a series of reactions of the type : 



RNHsOH + HCl = RNH 2 HC1 + H 2 O 



then we would obviously have to assume the existence of no fewer than 

 nine free NH 2 groups in the molecule of edestin. If we assume the 

 insoluble compound to have been formed by the union of two molecules 

 of acid with one of edestin, then the estimate of the number of free 

 NH 2 groups must be raised to 2X9= 18 and so forth. 



1 That is, the combining weight of HCl expressed in grams and multipled by 0.00014, 

 or the hydrochloric acid which is present in 1.4 c.c. of a tenth normal solution. 



