L.H.H* Duysens 



equal to the difference of the absorption spectra of oxidized 

 and reduced cytochrome, this shows that upon illumination a cy- 

 tochrome becomes oxidized. Similarly, if the fluorescence dif- 

 ference spectrum is equal to the fluorescence spectrum of chlo- 

 rophyll a, this means that the chlorophyll a fluorescence in- 

 creases upon illumination. Difference spectra determined between 

 various time limits make it possible in principle to analyze 

 the time course of photo synthetic intermediates which show ab- 

 sorption or fluorescence changes upon illumination. 



By measuring the effectivity of the actinic light as a func- 

 tion of the wavelength, it is possible in principle to identi- 

 fy the pigments which are responsible for the photochemical re- 

 action or reactions which cause changes in the absorption spec- 

 trum or the fluorescence spectrum. If these changes are quali- 

 tatively different in light of different actinic wavelengths, 

 then it follows that more than one photochemical reaction dri- 

 ven by pigment systems with different absorption spectra occurs. 

 This is the case in oxygen evolving photo synthesizing organisms. 

 In fluorescence experiments one more parameter is available: 

 the wavelength of the exciting light. The action or effective- 

 ness spectrum for exciting the fluorescence of a certain com- 

 pound is proportional to the sum of the absorption spectra of 

 the fluorescing substance and of the absorption spectra of 

 other pigments multiplied by the efficiences of transfer of ex- 

 citatien energy from these pigments to the fluorescing substan- 

 ce • 



Since it is possible to bring about oxidation or reduction 

 of a great number of non-physiological substances by means of 

 extracts of bacteria or higher plants, it is in general not 

 possible to conclude from these experiments, whether a certain 

 redox reaction occurs in the living cell, even if the reacting 

 substances are known to occur in the cell. For this reason 

 mainly, we have concentrated on studying reactions in intact 

 cells in order to establish which reactions occur i^ vivo . 

 Most if not all reactions observed by absorption and fluores- 

 cence techniques in living cells were redox reactions. In order 

 to find out whether a reaction is a main pathway in photosyn- 

 thesis or a side reaction, the quantum efficiency of this re- 

 action may be determined. Since the quantum requirement per 

 hydrogen atom or electron transported for COp reduction is 2, 

 an efficiency of the same order of magnitude may be expected 

 for the redox reactions in the photo synthetic chain. 



