280 Helmut Holzer 



For the explanation of the fructose diphosphate maximum a 

 surplus phosphorylation of fructose-6-phosphate can be ex- 

 cluded because of the behaviour of the ATP concentration; 

 the course of the ATP concentration is not at all correlated 

 with that of the fructose diphosphate concentration, as is 

 shown by Fig. 2. 



Furthermore, the behaviour of the orthophosphate concen- 

 tration shows that this substance, too, cannot have any share 

 in the formation of a fructose diphosphate maximum by 

 influencing the rate of triose phosphate dehydrogenation. In 

 order to explain the fructose diphosphate curve, within the 

 first 70 seconds a low and rate-limiting orthophosphate con- 

 centration, and after 70 seconds a high concentration of 

 orthophosphate should be expected, thus promoting the de- 

 gradation of fructose diphosphate via triose phosphate de- 

 hydrogenation. The observed behaviour of orthophosphate, 

 however, is in complete contradiction to this. 



A regulatory action of ADP upon triose phosphate dehydro- 

 genation can be excluded for the reasons mentioned above 

 with respect to ATP : the kinetics of the concentration changes 

 of this substance are by no means correlated with the kinetics 

 of the concentration changes of fructose diphosphate. 



As a last possibility we have examined whether a lack of 

 oxidized diphosphopyridine nucleotide (DPN), which is neces- 

 sary for triose phosphate dehydrogenation, might be respons- 

 ible for the accumulation of triose phosphate and fructose 

 diphosphate. During the first minute after glucose addition to 

 a yeast suspension, the concentration of acetaldehyde is so 

 small that it cannot be determined with our usual methods 

 (spectrophotometric test according to Warburg). This de- 

 pends upon the fact that acetaldehyde is in equilibrium with 

 alcohol, such that under anaerobic conditions only 1/200 of 

 the alcohol concentration is present as acetaldehyde (Holzer, 

 Holzer and Schultz, 1955). Moreover, the acetaldehyde 

 formed within the cells diffuses rapidly into the medium, and 

 for this reason cannot be concentrated in the cells in the same 

 way as the phosphorylated metabolites (Holzer, Schultz and 



