48 THE GENERAL CHARACTERS OF THE PROTEINS 



neutral reaction to litmus ; the preparations required from 0*85 to 1-5 

 c.c. to produce a neutral reaction to phenol phthalein when i gram of 

 the protein was employed. The causes of this acidity were investigated 

 by treating suspensions of known quantities of edestin with potassium 



/N\ 



hydroxide ( j solution until neutral with phenol phthalein, filter- 

 ing off the undissolved protein and certain quantities of protein pre- 

 cipitated by the addition of alkali (as the edestin crystals are partially 

 soluble in water, as will be explained later), and estimating quantita- 

 tively the constituents of the filtrate. These consisted of a certain 

 amount of organic matter together with potassium and sodium salts 

 of hydrochloric and sulphuric acids. The following conclusions 

 were drawn by Osborne as to the nature of edestin crystals : 



(1) That edestin is a basic body which forms true salts with acids. 



(2) That the preparations, as obtained by crystallisation from 

 salt solutions, react either weakly acid or neutral to litmus and are 

 salts of a basic protein. 



(3) That by the addition of caustic alkalis to these preparations 

 until the mixture no longer acts acid towards phenol phthalein, these 

 salts can be decomposed ; by the analysis of the filtrate from the pro- 

 tein after this treatment the nature of the salts can be determined. 



(4) The edestin preparations obtained by recrystallisation from 

 sodium chloride consist principally of chlorides (as the filtrate after 

 neutralisation with alkali contains chiefly chlorides) ; if prepared by 

 crystallisation from ammonium sulphate they consist chiefly of 

 sulphates. 



(5) The edestin preparations, as ordinarily prepared by crystal- 

 lisation from sodium chloride, are appreciably soluble in water. The 

 part which is soluble in water requires twice as much alkali for 

 neutralisation, with phenol phthalein as indicator, as the part which 

 is insoluble. 



(6) If the molecular weight of edestin be taken as 14,500, or 

 twice the simplest molecular weight as determined by direct analysis 

 (i.e., from C . H . N . S and O content), then the amount of hydrochloric 

 acid combined in the insoluble portion would correspond to a mono- 

 hydrochloride, whereas that in the soluble portion would correspond 

 to a dihydrochloride. The ordinary edestin preparations are, there- 

 fore, usually mixtures of chlorides, containing, however, a certain 

 quantity of sulphate. The relative amounts of these salts depend 

 on the relative amounts of chloride and sulphate present in the 

 liquids used either in the original extractions or in the recrystallisa- 

 tions. 



It is of interest to note that all the edestin preparations, whether 

 free base, sulphate or chloride, have the same crystalline form. The 

 protein part of the molecule is so large in comparison with that of 

 the combined acid that the latter exerts no influence on the form of 

 the crystals. Analogous instances of isomorphism are known in the 

 case of certain minerals, and also in the case of haemoglobin and its 

 derivatives (e.g., O and CO haemoglobin). In the above-mentioned 

 experiments the method of direct neutralisation was employed (me- 

 thod B I.) ; another series was carried out with the object of de- 

 termining the amount of the free base edestin (which is insoluble 

 in water) which is soluble in acids of given strength (method B II.). 



