160 PRINCIPLES OF GENERAL PHYSIOLOGY 



other ion does not pass through the membrane, it is better to take salts 

 of which the molecular weight and chemical constitution are known, since 

 quantitative results are easily obtained. Many of the aniline dyes with large 

 molecular weight answer this requirement. In the work done by myself 

 (1909, 1911), Congo-red and related dyes were found the most useful. It is 

 necessary to devote some consideration to this question, since the conditions 

 are rather complex, but salts of this nature are of frequent occurrence in the 

 cell, and play an important part therein. 



The exact chemical constitution of Congo-red is not material for the present 

 purpose, except that the coloured ion is the anion and, being a substituted di- 

 sulphonic acid, combines with two sodium ions. The anion is, of course, the one 

 to which the parchment paper membrane is impermeable. Measurements of the 

 electrical conductivity of these dyes show them to be electrolytically dissociated to 

 a considerable degree, so that the question to be answered is whether the sodium 

 ions are active osmotically when the membrane used is permeable to them. It 

 might indeed be supposed that these ions would pass through the membrane to 

 such a distance that their osmotic pressure was balanced by the electrostatic 

 attraction of the non-diffusible ions within the membrane, and that this fact would 

 render ineffective any pressure due to the kinetic energy of these ions on the 

 opposite side of the membrane. It must be confessed that the conditions are 

 difficult to grasp in thought, but it will be remembered that the osmotic pressure 

 produced by the non-diffusible substance, consisting of the anions and the non- 

 dissociated part of the salt, shows itself in virtue of the mechanical constraint 

 exerted by the membrane, which allows water to pass through freely, while holding 

 back the substances named. In a similar way, the sodium ions are held back by 

 the attraction of the opposite ions, which themselves are held back by the 

 membrane, so that the membrane itself must actually bear the pressure of both 

 kinds of ions. Or, to put it in another way, the pull of the anions on the cations 

 could not be effective unless the constraint of the membrane gave the former a 

 support to pull against. 



Experimental evidence, in any case, shows that all the ions actually present 

 are osmotically active. Vapour pressure measurements made by the method of 

 Barger, described above (page 155), gave the same values as direct measurements 

 with a parchment paper osmometer (Bayliss, 1911, p. 233). Now this vapour 

 pressure method gives the total molar concentration of the solution, including that 

 of the sodium ions, and therefore the parchment paper membrane does so also. A 

 still simpler proof that the diffusible ions are really active, is to take the dye, 

 " Chicago blue," in which the anion, like that of Congo-red, is a complex sulphonic 

 acid, but in this case there are four sulphonic acid groups in the molecule, so that 

 it combines with four sodium ions. If the latter were inactive, the osmot it- 

 pressure with a parchment paper membrane would be the same as that of an equally 

 concentrated solution of Congo-red, since the concentration of the non-diffusible 

 anion is identical ; in point of fact it is found to be double, hence the sodium ions 

 play their part. 



The matter is, however, not quite so simple. Although all the ions that are 

 present must be osmotically active, the numerical values of the osmotic pressure, 

 whether measured directly or by vapour pressure, are much less than would be the 

 case if the dissociation were of the usual simple kind of such an inorganic salt as 

 sodium chloride. The reason for this has not yet been satisfactorily made out, but 

 there seems to be no doubt that it depends on the formation of complex, aggregated 

 ions. The remarkable fact is that electrical conductivity measurements give no 

 evidence of a less total number of charges than if no aggregation existed. The 

 complex ions appear to possess the same number of charges as if their constituents 

 were free. 



The way in which the electrostatic forces at the membrane influence the 

 distribution of diffusible saltc between the two sides of the membrane has been 

 referred to above (page 120). It is found that, suppose the dye is a sodium salt, and 

 the diffusible salt is sodium chloride, the distribution is such that there is always 

 less of the sodium chloride within than without. The explanation is, no doubt, 



