KLUYVER S CONTRIBUTIONS TO MICROBIOLOGY AND BIOCHEMISTRY 



participating in metabolism may often cause lower potentials than 

 those measured by means of the metal electrodes in the cell suspensions.' 



In this manner the apparent contradiction in the results obtained 

 by Kluyver and Hoogerheide and by Fromageot and Desnuelle, re- 

 spectively, could readily be explained. It was but necessary to assume 

 that the dyes could interact with intracellular catalysts, representing 

 a redox system at an even lower potential, but not excreted into the 

 medium, and therefore unable to exert an effect on the electrodes. 

 Consequently the potentials established at the electrodes did not 

 necessarily represent the potentials inside the cells resulting from the 

 particular metabolic activity under investigation. At the same time, 

 the observed reduction of Nile blue suggested to Kluyver and Hooger- 

 heide a simple method whereby the potential in the interior of metab- 

 olizing cells might become measurable with the aid of an external 

 electrode. This consisted in the addition to the cell suspension of a 

 readily diffusible redox dye of appropriate normal potential; it was 

 felt that the dye would mediate the necessary contact between the 

 intracellular catalysts and the electrodes. A series of measurements 

 conducted in this manner showed that now the electrode potentials 

 in fermenting yeast suspensions were indeed much lower than those 

 previously determined, and fell in the range of —30 to —40 mV. It 

 is true that some exceptions were noted; in suspensions containing 

 certain dyes with normal potentials high enough to warrant the ex- 

 pectation that they should at least be partially reduced - e.g. indigo 

 di-, tri-, and tetra sulphonates - the measured potentials were once 

 again found to be around 100 mV. This result could, however, be 

 attributed to an inability of these substances to penetrate into the 

 cells, an interpretation that was supported by tests showing that 

 such indicators are not reduced at all. 



These experiments led Kluyver and Hoogerheide to introduce a 

 general technique for the measurement of redox potentials free from 

 the ambiguity resulting from a dependency on excreted redox sys- 

 tems. The technique involves measurements of electrode potentials in 

 cell suspensions supplemented with a mixture of redox dyes covering 

 a wide range of normal potentials, a 'universal indicator mixture'. 

 Appropriate experiments with the various yeasts and Ps. lindneri then 

 revealed that under these conditions an electrode potential of —30 to 

 —40 mV was established in suspensions of all of them. This was therefore 



