Abstract
Rapid microdetermination of D-glucose anomers with β-D-glucose dehydrogenase and mutarotase was deviced. The reaction mixture (1ml) composed of 20 units of purified β-D-glucose dehydrogenase (Bacillus cereus M 1020) and 2μmole of NAD+ dissolved in 20mM EDTA buffer (pH 6.5). By addition of D-glucose solution (5μl,<30μg of D-glucose) at 25°, arapid increase of absorption (NADH) at 340nm due to β-D-glucose was recorded, then 5units of mutarotase (hog kidney) was added in the reaction mixture to mutarotate the remaining α-D-glucose to β-D-glucose. The second increase of absorption (NADH) due to the α-D-glucose was recorded. From these two absorptions, the ratio of β to α-D-glucose was determined in 3.5-4min. This new method was not interfered by glutathione, cysteine, ascorbic acid, sodium fluoride, uric acid, etc. β-D-Glucose dehydrogenase from Gluconobacter oxydans can be used also for the same determination of D-glucose anomers, however, NADP should be used as a coenzyme
