ACTIVATION OF FLUORESCENCE 769 



nonactivated one, is independent of the nature of the activator, and bears 

 no relation to the absorption spectrum of chlorophyll in pure activator. 



The fluorescence of activated solutions is quenched by rising tempera- 

 ture (15-70° C), particularly strongly in the region of partial activation. 

 The maximum intensity, Fo, is a linear function of temperature over a 

 comparatively wide range (a similar relationship was found also by Zscheile 

 and Harris, 1943). In pai'tly activated solutions, the temperature curve 

 is not only steeper than in fully activated ones, but also shows — with some 

 activators at least — a definite curvature. 



The concentration of the strongest known activators, required to achieve 

 complete activation, is of the same order of magnitude as that of chloro- 

 phyll itself, which in the experiments of Livingston et al. was 5 X 10 ~^ 

 mole/1. This shows that the effect cannot be due to kinetic encounters 

 between chlorophyll and activator (which are too rare at such low concen- 

 trations) ; nor can it be ascribed to a change in properties (such as dielec- 

 tric constant) of the solvent as a whole. Rather, the effect must be caused 

 by the association of chlorophyll molecules with the molecules of the activa- 

 tor. The change in absorption spectrum supports this assumption. If the 

 nonassociated form is totally nonfluorescent, the intensity of fluorescence 

 can be used to calculate the proportion of chlorophyll molecules in the as- 

 sociated form — assuming that the absorption coefficient of associated 

 chlorophyll is the same as that of the nonassociated pigment. Figvire 

 21. 26. A shows that this is not quite true for chlorophyll a at 436 ni/x; but 

 Livingston neglected this difference. Assuming a one-to-one complex 

 [ChlAc ] the equilibrium constant of association : 



can be calculated from the half-activating concentration [Ac]i/,: 



K[Chl]o[Ac]o 



(23.4B) [ChlAc] = 



1 + K[Ac]o 



^^^•■*^^ Fo " 1 + A'[Ac]o 



(23.4D) A' = ^ 



Ac],/, 



{cf. chapter 27, eq. 27.12). This is a simplified solution, based on the as- 

 sumption [Ac] ^^ [Ac]o; in other words, it assumes that the amount of 

 activator bound in the complex is small compared to the total amount 

 added to obtain activation. In Table 23.IIIA, the concentration [AcChl] 

 at half-activation is 2.5 X 10 ~^ mole/1, for piperidine; since [Ac Jo = 

 6.5 X 10-^ [Ac]i/, = 4 X 10"" molc/1. In other words, the simplified 

 equations are not quite applicable to piperidine (and four other systems 



