Nature of the Acceleration Produced by Arsenate and Arsenite.
In explanation of the remarkable accelerating action of arsenates and arsenites two obvious possibilities present themselves. In the [p079] first place the arsenic compound may actually replace phosphate in the reaction characteristic of alcoholic fermentation, the resulting arsenic analogue of the hexosephosphate being so unstable that it undergoes immediate hydrolysis, and is therefore only present in extremely small concentration at any period of the fermentation and cannot be isolated. In the second place it is possible that the arsenic compound may accelerate the action of the hexosephosphatase of the juice, and thus by increasing the rate of circulation of the phosphate produce the permanent rise of rate. With this effect may possibly be associated a direct acceleration of the action of the fermenting complex.
The experimental decision between these alternative explanations is rendered possible by the use of a mixture of enzyme and co-enzyme free from phosphate and hexosephosphate. As has already been described (p. [55]) a mixture of boiled yeast-juice, which has been treated with lead acetate, glucose or fructose, and washed zymin can be prepared which scarcely undergoes any fermentation unless phosphate be added. If now arsenates or arsenites can replace phosphate, they should be capable of setting up fermentation in such a mixture. Experiment shows that they do not possess this power. For fermentation to proceed phosphate must be present and it cannot be replaced either by arsenate or arsenite [Harden and Young, [1911, 1]].
The effect of these salts on the action of the hexosephosphatase can also be ascertained by a modification of the foregoing experiment. If a hexosephosphate be made the sole source of phosphate in such a mixture as that described above, in which it must be remembered abundance of sugar is present, the rate at which fermentation can proceed will be controlled by the rate at which the hexosephosphate is decomposed with formation of phosphate. Experiment shows that in the presence of added arsenate or arsenite the rate of fermentation is largely increased, so that the effect of these salts must be to increase the rate of liberation of phosphate, or in other words, to accelerate the hydrolytic action of the hexosephosphatase.
This conclusion is even more strikingly confirmed by a comparison of the direct action of yeast-juice on hexosephosphate in presence and in absence of arsenate, as measured by the actual production of free phosphate. In a particular experiment this gave rise to 0·0707 gram of Mg2P2O7 in the absence of arsenate and 0·6136 gram of Mg2P2O7 in the presence of arsenate.
The results obtained with arsenite are throughout very similar to those given by arsenate, but are not quite so striking. It may therefore be affirmed with some confidence that the chief action of arsenates [p080] and arsenites in accelerating the rate of fermentation of sugars by yeast-juice or zymin, consists in an acceleration of the rate at which phosphate is produced from the hexosephosphate by the action of the hexosephosphatase.
It has further been found that arsenates, and to a less degree arsenites, also produce an acceleration of the rate of autofermentation of yeast-juice and of the rate at which glycogen is fermented. This turns out to be due in all probability to an increase in the activity of the glycogenase by the action of which the sugar is supplied which is the direct subject of fermentation. Thus in one case an initial rate of fermentation of glycogen of 1·9 c.c. per five minutes was increased by 0·05 molar arsenate to 9·7 and the amount of carbon dioxide evolved in two hours from 38 to 158 c.c. Even this enhanced production of glucose from glycogen, however, is not nearly sufficient for the complete utilisation of the phosphate also being liberated by the action on the hexosephosphatase, for the addition of an excess of sugar produces a much higher rate, in this case 36 c.c. per five minutes. The effect of arsenate on the rate of action of the glycogenase seems therefore to be much smaller than on that of the hexosephosphatase.
No other substances have yet been found which share these interesting properties with arsenates and arsenites, and no advance has been made towards an understanding of the mechanism of the accelerating action of these salts on the specific enzymes which are affected by them.