Evaluation of Glucose-6-Phosphate Dehydrogenase from Leuconostoc mesenteroides in the Hexokinase Method for Determining Glucose in Serum

Abstract

The hexokinase (EC 2.7.1.1)/glucose-6-phosphate dehydrogenase (I) (EC 1.1.1.49) method of glucose analysis is highly accurate, precise, and sensitive when I from Leuconostoc mesenteroides and NAD+ are used. Coefficients of variation ranged from 0.68 to 8.9% for glucose standards between 0.56 and 27.7 mmol/liter. Recovery was 103 ± 10% for glucose standards added to sera of known glucose content. Although no substances investigated interfere with the test system when present at their normal or likely concentrations in serum, final reaction mixtures containing mannose (>0.28 mmol/liter), fructose (3.7 mmol/liter), or 2-deoxy-D-glucose (4.01 mmol/liter) inhibit the analysis. With all constituents lyophilized, the reagent mixture is stable for at least a year at 4 °C and for five days at 40 °C. Within-day reproducibility averaged 0.83%; day-to-day precision was 1.3%. We compared results of hexokinase/glucose procedures by using I from yeast (with NADP+) and from L. mesenteroides (with either NAD+ or NADP+ as coenzyme); there was complete correlation over a serum glucose range of 4 to 12 mmol/liter (72 to 217 mg/dl). Use of NAD+ with I from L. mesenteroides is advantageous because of the decreased cost and greater stability of the coenzyme.