STUDIES OH PRIMARY REACTIOllS AUD HYDROGEN OR ELECTRON TRANSPORT 

 IN PHOTOSYHTHESIS BY MEANS OP ABSORPTION AND FLUORESCENCE DIF- 

 FERENCE SPECTROPHOTOMETRY OP INTACT CELLS 



L.N.M. Duysens 



INTRODUCTION AND METHODS 



As requested by the organizers of this symposium, in this 

 paper investigations mainly carried out at our laboratory will 

 be reviewed and interpreted. 



In most experiments the time courses of changes in light ab- 

 sorption and emission were recorded at various wavelengths in 

 the visible and adjacent spectral regions for suspensions of in- 

 tact photo synthesizing cells. These changes were brought about 

 by suddenly admitting relatively strong so-called actinic light 

 of constant intensity. This light was switched off after a time 

 varying from one second to about one minute. The changes in ab- 

 sorption were measured by means of a separate weak modulated 

 measuring, beam, which in general did not cause changes in ab- 

 sorption ' J the fluorescence was in general excited by modu- 

 lated light of such a/^Qw intensity so as not to cause changes 

 in fluorescence yield^ . The measuring apparatus was only sen- 

 sitive to the modulated measuring beam or to the fluorescence, 

 excited by the modulated exciting beam. These modulations and 

 the use of suitable filters prevented direct effects of the ac- 

 tinic light on the measuring apparatus. The absorption and fluor- 

 escence changes were recorded by means of a one-fourth or one 

 second recorder. Rapid changes in fluorescence yield were brought 

 about by an electronic flash, and were measured by means of an 

 oscillograph. This apparatus has a response time of the order 

 of one usee, and the flash served both as exciting and actinic 

 beam • 



A difference spectrum presents a change in the absorption 

 spectrum or/Of the fluorescence spectrum occurring during il- 

 lumination » Difference spectra may be obtained from the ti- 

 me c\irves of absorbancy or fluorescence changes (see ). These 

 spectra sometimes permit the identification of substances cau- 

 sing the spectral changes. If e.g. the difference spectrum is 



