168 LIGHT AND LIFE 



free radicals which are measurable by EPR. It is therefore clear 

 that no simple monomeric semiquinone is formed under such con- 

 ditions. A more complicated situation obtains. In those cases in 

 which a persistent band is observed, a very strong binding of the 

 substrate to the enzyme occurs (17) . Interaction with neighboring 

 substrate radicals in such a complex may make semiquinones vmde- 

 tectable by EPR. Since many flavoproteins contain two prosthetic 

 flavins, flavin biradical formation may also occur in these complexes. 

 EPR signals would not be expected in this case. The spectral band, 

 however, which is clue to transitions of a considerably higher energy 

 level would not necessarily be influenced by such interactions and 

 would thus still be visible. 



It is not easy to prove that interactions of the type mentioned are 

 indeed responsible for the failure of EPR to detect free radicals when 

 certain flavoproteins react with substrate and Avhen the spectral band 

 appears which was found characteristic of semiquinone formation in 

 other cases. One may have to consider other possibilities of explaining 

 the observations. It appears extremely unlikely that the strong com- 

 plexes formed between what was initially oxidized flavoprotein and 

 reduced form of substrate would not include some kind of oxido- 

 reductive interaction. It has been suggested that this interaction may 

 be of a charge transfer type. This contention again is hard to prove 

 experimentally. Absence of EPR signals is certainly in accord with 

 it, but the fact that the characteristic absorption band is found 

 in almost the same position with a variety of flavoproteins and sub- 

 strates is not in agreement with observations on charge transfer 

 bands (12) . The occurrence of a charge transfer absorption also im- 

 plies that charge transfer only takes place significantly in the excited 

 state. One may therefore wonder whether an enzymatic oxidoreduc- 

 tion reaction could proceed efficiently between a substrate and an 

 enzyme which require energy of the magnitude of light energy for 

 electron transfer. 



It is possible that present concepts and terminology are not able 

 to describe satisfactorily the kind of intermediate and of interaction 

 which we are actually dealing with in these complexes. 



Although I have tried in the foregoing remarks to arrive at a 

 few general conclusions from a vast and diverse body of experimental 

 material, I would at the same time like to express my view that 

 generalizations are hazardous and that at this early stage of our 

 knowledge of the phenomena imder discussion we have little choice 

 but to consider every case individually and to study it on its own 

 merits. 



