788 FLUORESCENCE OF PIGMENTS IN VITRO CHAP. 23 



can be produced also by small amounts of salts, such as LaCls or CeCls in 

 the presence of air. These salts probably catalyze the allomerization of 

 chlorophyll by air, with the concomitant changes in the intensity of fluores- 

 cence (decrease with chlorophyll a, increase with chlorophyll h). These 

 catalyzed reactions, too, occur only in alcoholic solutions. In ethereal 

 solution, chlorophyll a is converted by LaCls into a yellow compound, and 

 all fluorescence soon vanishes. 



Evstigneev and Krasnovsky (1948) and Evstigneev, Gavrilova and 

 Krasnovsky (1949^) also have investigated the quenching of chlorophyll 

 fluorescence by several substances (see footnotes to Table 23. HID), in 

 particular by oxygen. Their peculiar initial result already was mentioned 

 on page 648: they found the effect of oxygen to be strongly dependent 

 on the solvent. In polar solvents — pyridine, ethanol, ethyl acetate, 

 acetone (and also in commercial benzene) — the effect of oxygen was as 

 previously described: moderately strong quenching. In nonpolar sol- 

 vents — heptane, toluene, carbon tetrachloride — on the other hand, an 

 entirely different effect was found; fluorescence (of chlorophyll a or a + 

 b) decreased upon removal of oxygen (evacuation of the vessel by an oil 

 pump) by as much as a factor of two; it increased to approximately the 

 original level after readmission of air. (The fact that benzene behaved 

 like a polar solvent probably was due to an impurity.) Repeated evacua- 

 tion and aeration produced a gradually weakening effect — a result which 

 could be attributed to superposition, upon reversible association of chloro- 

 phyll with oxygen, of an irreversible oxidation (allomerization). Later 

 (1949^), the same investigators reported that Avhat they first took for an 

 activating effect of oxygen was in fact an activating effect of water vapor , 

 contained in the admitted air. Admission of air caused no activation of 

 fluorescence in moist solvents. As described on p. 771, Livingston at- 

 tributed activation to an enol-ketone transformation in the cyclopentanonc 

 ring, enhanced by the formation of hydrogen bonds, while Evstigneev and 

 co-workers thought that polar solvent molecules might have an affinity 

 for the magnesium atom (which, in the chlorophyll molecule, has two free 

 coordination places). Saturation of these affinities could stiffen the 

 molecule and delay internal dissipation of excitation energy. In agree- 

 ment with this concept, no activation was obtained with Mg-free pheo- 

 phytin, or phthalocyanin {cf. p. 772). 



Coe (1941) suggested that "chemical" quenching of chlorophyll fluorescence by 

 "antioxidants" present in oils and fats, could be used practically as a measure of their 

 rancidity; but French and Lundberg (1944) found no evidence of such quenching by 

 cottonseed oil. 



Kautsky, Hirsch and Flesch (1935) found, and Franck and Livingston 

 (1941) confirmed, that certain substrates whose photoxidation is sensitized 



