FLUORESCENCE OF PIGMENTS in vivO 1867 



Sharp scattering peaks were found to correspond not only to the main 

 chlorophyll bands, but also to the carotenoid bands, almost hidden in the 

 absorption spectrum. Selective scattering was noted also in colloidal 

 chlorophyll suspensions, indicating that it does not require a regular pig- 

 ment lattice, but merely a sufficiently close packing of pigment molecules. 



The occurrence of sharp selective scattering bands poses additional 

 problems for the application of scattering correction to absorption meas- 

 urements in vivo. On the other hand, it opens a new approach to the eluci- 

 dation of the physical state of the pigments in the living cell ; for example, 

 the sharpness of the carotenoid bands in scattered light can be taken as an 

 indication of a particularly close (or particularly regular) arrangement of 

 their molecules (as compared to those of the other pigments). 



Using an integrating sphere assures one against including scattering 

 in absorption; but it does not prevent absorption from being itself affected 

 by scattering, through increased pathway of the light in the absorbing me- 

 dium ("detour factor"). 



7. Fluorescence of Pigments in vivo 

 (Addendum to Chapter 24) 



(a) Absolute Yield 



Duysens (1952, page 84 and page 88) made criticisms of the estimates 

 of the yield of chlorophyll and bacteriochlorophyil fluorescence in vivo, 

 which led Wassink et al. to figiu'es of the order of 0.15%, for both Chlorella 

 and Chromatium, while yields of ^1.5% were estimated for Chroococcus 

 by Arnold and Oppenheimer (c/. chapter 24, section 2, and chapter 32, 

 section 6). Duysens suggested that neglected geometrical-optical factors 

 have reduced the observed yield in green algae and bacteria by a factor 

 of perhaps 2.7, and that re-absorption of fluorescence accounted for a loss 

 of another 50%; so that the true quantum yield of fluorescence in these 

 cells may be of the order of 1% rather that 0.1%. Obviously, this criticism 

 calls for careful experimental re-examination. Preliminary measurements 

 by Latimer (unpublished) seem to support it (cf. page 1992). 



Duysens further suggested that, conceivably, the "intrinsic" fluo- 

 rescence capacity of chlorophyll (or bacteriochlorophyil) in the natural 

 state is another order of magnitude higher (i. e. of the order of 10%, as 

 in vitro), and that the reduction of the actual fluorescence to about 1% 

 may be due to a 90% effective resonance energy migration to "reduction 

 centers," where this energy is used for a photochemical transformation 

 (cf. above section 3). Duysens estimated that this is possible if about 

 200 energy transfers are needed to reach a "center." 



We can obtain a similar, but somewhat larger estimate, by the crude 



