I86G SPECTROSCOPY AND FLUORESCENCE OF PIGMENTS CHAP. 37C 



the suspension encounters several particles, and therefore must be in- 

 significant in the usual suspensions of algae, were incorrect; the effect 

 remains significant even in ''dense" Chlorella suspensions (containing per- 

 haps 10^ cells, with a total volume of the order of 10~'* cc, in each cubic 

 centimeter of the liquid). Only when the volume of cells reaches the 

 same order of magnitude as the total volume of the suspension, does 

 relation 37C.17 cease to be valid. These considerations should be taken 

 into account in attempts to analyze tiansmission curves of cell systems in 

 terms of the absorption curves of the several pigments as observed in 

 extracts. In addition to band shift, one must consider band flattening by 

 the sieve and bunching effects, before concluding that intrinsic changes in 

 the absorption band shape have occurred. It will be noted that, for a 

 particle transmission of 0.5, the ratio: 



(1 - T,)/ In (l/Dp 



is 0.5/0.7, or 70%; for a transmission of 0.1, it is 0.9/2.3, or close to 40%. 

 This shows how strong the flattening effect is if single particles have a 

 high optical density. 



Applying the band flattening theory to Chlorella absorption at 680 m^, 

 Duysens found 1 — Tp = 0.63, in agreement with the figure (64%) derived 

 above from the asymmetry of fluorescence. 



6A. Scattering by Pigment Bodies in vivo 



All the above considerations did not take into account scattering, which 

 can complicate the interpretation of absorption measurements on sus- 

 pensions. Observations of Jacobs et al. on suspensions of chlorophyllide 

 microcrystals (section 3) showed that, in addition to the "Rayleigh scat- 

 tering," decreasing steadily from the shorter to the longer waves, one may 

 have to consider selective scattering, with a peak on the long-wave side 

 of the absorption band, dropping rapidly on the short-wave side and de- 

 clining much more slowly on the long-wave side. The puzzle of the 

 increased "absorption" of live cells and crude aqueous cell dispersions in 

 the far red, noted on p. 715, may have its answer in such selective scattering. 



In the absence of direct evidence it could be argued that, in vivo, pig- 

 ments will not produce selective scattering, even if they are arranged in 

 complete monolayers, because scattering requires an extension of the regu- 

 lar arrangement in all three dimensions. Duysens (1954) suggested, in 

 fact, that a correction for Rayleigh scattering is adequate in this case, 

 to take care of excessive apparent absorption in the far red as well as in the 

 green. 



However, preliminary experiments by Latimer (1954) have revealed a 

 very pronounced selective scattering of light by Chlorella suspensions. 



