Spectrophotometric measurement of hexokinase and phosphohexokinase activity.

The methods available for the quantitative assay of these enzymes are based on phosphate transfer determinations and pH change of the reaction mixture. They require large quantities of adenosine triphosphate and, for estimation in crude tissue extracts, the addition of inhibitors such as iodoacetate and sodium fluoride (7). The purpose of this paper is to report a simple and rapid spectrophotometric method by which the rates of phosphorylation by the enzymes hexokinase and phosphohexokinase can be measured. Recently, spectrophotometric tests for the other principal enzymes of the glycolytic system of carbohydrate breakdown have been developed, mainly by Warburg and his collaborators. These methods should prove useful tools in the study of disturbances of carbohydrate metabolism, such as those described for tumor tissues. It should be emphasized, however, that several of them are not suitable for crude tissue extracts and considerable modification may prove necessary to adapt the methods for such purposes.