Methods for Measurement and Analysis of Changes 



in Light Absorption Occurring upon Illumination 



of Photosynthesizing Organisms 



L. N. M. DUYSENS, Biophysical Research Group, Department of Physics, 

 University of Utrecht, Utrecht, Netherlands 



Upon illumination of photosynthesizing cells a change will occur 

 in the concentration of all intermediates and catalysts participating 

 in photosynthesis. If the changes in concentration result in measur- 

 able variations in absorption, these variations at various wave- 

 lengths provide information about the kind and function of inter- 

 mediates and catalysts in photosynthesis. 



For colorless cells and their extracts, measurements of spectral 

 changes in absorption of intermediates and catalysts brought about 

 by adding substrates, have proved of utmost importance for the 

 elucidation of biochemical reactions. The magnitude of the change in 

 optical density is proportional to the concentration of cell material. 

 Fairly dense cell suspensions have been used in most experiments. 

 It is not possible, in general, to use concentrations of photosynthe- 

 sizing cells as high as those of colorless cells, because the photosyn- 

 thesizing cells are so strongly colored that they would absorb prac- 

 tically all the measuring Hght. Since the concentration of substances 

 changing their absorption seems to be of the same order of magnitude 

 in photosynthesizing and colorless cells, this may be a reason why, for 

 measurements on photosynthesizing cells, generally an apparatus is 

 required ten to a hundred times more sensitive than the apparatus 

 usable for "colorless" cells. In addition precautions should be taken 

 that the light used for bringing about a change in absorption does 

 not affect the measuring apparatus directly. 



APPARATUS AND METHODS FOR MEASURING ABSORPTION 



CHANGES 



The experimental problem is to measure only those variations in 

 intensity of the monochromatic light beam transmitted by a sus- 

 pension of photosynthesizing cells which are caused by changes in 



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