Publisher Summary This chapter discusses an enzymatic method for the determination of D -glucose with hexokinase and glucose-6-phosphate dehydrogenase. The optical method for the determination of glucose-6-phosphate, on which the method discussed in the chapter is based, was developed by O. Warburg and his collaborators. Hexokinase catalyzes the phosphorylation of glucose by adenosine triphosphate (ATP). Glucose-6-phosphate is oxidized in the presence of triphosphopyridine nucleotide (TPN) by glucose-6-phosphate dehydrogenase. For all practical purposes, both the reactions proceed stoichiometrically and quantitatively, although reversibility has been demonstrated under special conditions. The reduced triphosphopyridine nucleotide (TPNH) arising in the second reaction is determined spectrophotometrically at 340 or 366 mμ and serves as a measure of the glucose-6-phosphate formed from the glucose in the first reaction. The preparation need not be crystalline but must be relatively free from compounds that interfere with the glucose determination. Sometimes, the preparations still contain glucose, which has been added as a stabilizing agent during purification. It can be removed from highly purified hexokinase by dialysis. The least interference from contaminating enzymes occurs if the specific activity of the hexokinase preparation used approaches that of the crystalline enzyme.

d-Glucose: Determination with Hexokinase and Glucose-6-phosphate Dehydrogenase

(1960). On the Enzymatic Determination of Blood Glucose. Scandinavian Journal of Clinical and Laboratory Investigation: Vol. 12, No. 4, pp. 402-407.

On the Enzymatic Determination of Blood Glucose

Photometric method for the determination of low concentrations of hydrogen peroxide by the peroxidase catalyzed oxidation of N,N-diethyl-p-phenylenediamine (DPD)

Analysis of a functional change in membrane in the process of cell transformation by Rous sarcoma virus; alteration in the characteristics of sugar transport.

A methodological investigation of the glucose oxidase method for determination of glucose is described. The importance of eliminating substances reducing H2O2 generated from glucose and other substances causing unspecific oxidation of the chromogen is emphasized. On the basis of this study a method has been constructed which gives accurate glucose values in whole blood, plasma, and red blood cells.

J. E. Johnson

Papers

Enzymatic Microdetermination of Glucose in Blood and Urine