1 88 PRINCIPLES OF GENERAL PHYSIOLOGY 



always a certain proportion of the total acid present, so that the moment a part 

 of the acid has been removed by the addition of a base, the remaining acid 

 undergoes a further dissociation and so on, until the whole of the acid, whatever 

 its original dissociation was, has become completely dissociated and its hydrogen 

 ions have entered into combination with the hydroxyl ions of the base. 



There are, however, certain methods by which the actual hydrogen ion 

 concentration can be estimated without causing any change in it. 



We will first consider the use of Indicators. These are certain dyes which 

 have a particular colour at a certain concentration in H' ions and another colour 

 at another concentration which differs very little from the first. Those which 

 change colour at points not far distant from neutrality are the most useful, 

 especially in physiological work. 



That it is really the hydrogen ion concentration that these substances "indicate" is 

 obvious if we take a series of five dilutions of hydrochloric acid, viz. , twice normal, normal, 

 dci-, centi-, and milli-normal ; the colour of crystal violet will be found to be yellow in the 

 first, yellow-green in the second, blue-green in the third, blue in the fourth, and violet in 

 the fifth. No alkali has been added, and the only difference between the various solutions is 

 the concentration in the acid. 



The whole question of the theory of indicators cannot be entered into here, but may be 

 found in Nernst's book (1911, pp. 533-536). Generally speaking, they are salts of either 

 a very weak acid or a very weak base, sometimes the free acid or base itself. The change in 

 colour is due to the electrolytic dissociation of the salt with the production of an ion which 

 has a different colour from that of the free undissociated acid or base. 



Since the strength of the indicator acid or base varies in the different 

 substances used for the purpose, it will be clear that the acidity of a given 

 solution may be determined by the use of a series of indicators changing colour 

 at different H' ion concentrations. In theory, the question is a little complicated 

 by the existence of what are known as " pseudo-acids," which have a different 

 chemical structure in the free state to fliat in their electrolytically dissociated 

 salts ; but the explanation given, which was originally due to Ostwald, is not 

 practically altered by this fact. 



That indicators do actually vary in the acidity of the solution to which they respond can 

 easily be seen by comparing methyl orange with phenolphthalein. If a solution of hydro- 

 chloric acid be taken it will be found that methyl orange is red in it. Alkali is now added 

 until the colour changes to orange, that is, the solution is alkaline to this indicator. If another 

 sample of the acid be taken, it will be found to produce no colour with phenolphthalein, and 

 more alkali must be added to change the colour of this indicator to the red one of its salts 

 than was required to change the colour of methyl orange 



In the use of indicators there are several precautions to be observed. 



In the first place, the hydrogen ion concentration at which certain of them 

 change colour is not the same in pure acids or bases as in the presence of foreign 

 substances, especially salts and proteins. For a description of these cases, the 

 reader is referred to the investigations of Sorensen (1909), which are concerned 

 with the various methods of practical use for the estimation of hydrogen ion 

 concentrations. The use of indicators for physiological purposes will be found 

 fully treated. In the second place, it will be obvious that the total amount of the 

 indicator present must not be so great as to neutralise, or react with, any 

 perceptible portion of the ions to be estimated. 



This will be made clear if we take a dilute solution of Congo-red, the sodium salt of an acid 

 whose coloured ion is red and whose undissociated free acid is blue. Add a drop of this solu- 

 tion to a very dilute solution of hydrochloric acid, a blue colour is given. Take again a 

 concentrated solution of the indicator and add it in rather large amount to a small quantity of 

 the very dilute acid. The colour will remain red, because the whole of the free hydrochloric 

 acid present has been used up to combine with a portion only of the dye, and the colour of tl it- 

 salt still left in excess masks the bluish colour or the very small amount of the free dye-acid. 

 This fact is especially liable to mislead when test papers are used, and a drop of very dilute 

 solution, or one containing only a very small amount of hydrogen ions, is applied to the paper, 

 as has been pointed out by Walpole (1913, 1). In such cases the reaction will appear to 

 be different when a drop is placed on the paper and when the paper is immersed in a 

 large volume of the solution. 



Walpole (1910) has also described an ingenious artifice by which it is possible to use 

 an indicator with solutions containing coloured substances. This method consists essentially 

 in comparing the colour of the solution to which an indicator has been added with that of the 

 light which has first passed through an equal depth of the coloured solution alone, and 



